Andrographolide Analogues Research Articles

Discovery and development of labdane-oxindole hybrids as small-molecule inhibitors against chikungunya virus infection

Chikungunya virus (CHIKV) infection, a febrile illness caused by a mosquito-transmitted alphavirus, has afflicted millions of people worldwide. There is currently no approved effective antiviral treatment for CHIKV infection. In this study, we report a new class of small-molecule CHIKV inhibitors, the oxindole-labdanes, that potently block the replication of CHIKV with good selectivity. Andrographolide, a previously reported inhibitor of CHIKV infection, was used as the lead compound for our initial structure-activity relationship (SAR) study. From a focused library of 72 andrographolide analogues, we identified the lead compound (E)-2 with improved antiviral activities. Further optimization of (E)-2 led to the discovery of the normal-labdane 7-chloro-oxindole (E)-42 as potent inhibitor against two low-passage CHIKV isolates from human patients with an EC50 of 1.55 μM against CHIKV-122508 and 0.14 μM against CHIKV-6708. Compound (E)-42 displayed minimal cytotoxic liability (CC50 > 100 μM), thus furnishing good selectivity relative to the host cells. Mechanistically, (E)-42 does not inactivate the viral particles but rather acts on the host cells to interfere with the viral replication, demonstrating both prophylactic and therapeutic effects. Our findings open a new avenue for the development of oxindole-labdane compounds as promising antiviral drugs against CHIKV infection.

Reactivity-informed design, synthesis, and Michael addition kinetics of C-ring andrographolide analogs

Andrographolide, a labdane diterpenoid lactone natural product extracted from Andrographis paniculata, has demonstrated potent biological activity and therapeutic potential against cancer, Alzheimer's disease, diabetes, and multiple sclerosis. Andrographolide is reported to significantly inhibit the NF-κB signaling pathway, which is active in immune system function and regulation of inflammatory cells. However, andrographolide use is not optimized for human systems. In order to improve upon these aspects, we designed and semi- synthesized a library of andrographolide analogues with modified electronics of the 12,13 unsaturated lactone to increase cytotoxicity via the addition of electron-withdrawing groups, thereby influencing reactivity of the compound. Reactivity of the top piece butenolide warhead was quantified via a time- resolved colorimetric ex vivo Michael addition assay using Elleman’s Method, wherein reduced-glutathione was the electron donor. The results demonstrate that the installation of an acetate ester at C14 did not result in greater Michael acceptor reactivity, whereas changing the electrophile to C14 with a C11-C12 alkene resulted in a significantly higher rate of reaction with glutathione. Further, computational studies were employed to model the Michael addition energetic pathways of andrographolide and each analog. We found the energetic pathways for all analogs were relatively consistent, suggesting that changes in sterics may play a more significant role in defining Michael addition kinetics. Through understanding the kinetic mechanisms of the Michael acceptor pharmacophore in andrographolide, we hope to further inform directed semisyntheses in order to optimize this incredibly potent natural product for clinical use.

Discovery of C-12 dithiocarbamate andrographolide analogues as inhibitors of SARS-CoV-2 main protease: In vitro and in silico studies

A global crisis of coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has impacted millions of people’s lives throughout the world. In parallel to vaccine development, identifying potential antiviral agents against SARS-CoV-2 has become an urgent need to combat COVID-19. One of the most attractive drug targets for discovering anti-SARS-CoV-2 agents is the main protease (Mpro), which plays a pivotal role in the viral life cycle. This study aimed to elucidate a series of twenty-one 12-dithiocarbamate-14-deoxyandrographolide analogues as SARS-CoV-2 Mpro inhibitors using in vitro and in silico studies. These compounds were initially screened for the inhibitory activity toward SARS-CoV-2 Mpro by in vitro enzyme-based assay. We found that compounds 3k, 3l, 3m and 3t showed promising inhibitory activity against SARS-CoV-2 Mpro with >50% inhibition at 10 μM. Afterward, the binding mode of each compound in the active site of SARS-CoV-2 Mpro was explored by molecular docking. The optimum docked complexes were then chosen and subjected to molecular dynamic (MD) simulations. The MD results suggested that all studied complexes were stable along the simulation time, and most of the compounds could fit well with the SARS-CoV-2 Mpro active site, particularly at S1, S2 and S4 subsites. The per-residue decomposition free energy calculations indicated that the hot-spot residues essential for ligand binding were T25, H41, C44, S46, M49, C145, H163, M165, E166, L167, D187, R188, Q189 and T190. Therefore, the obtained information from the combined experimental and computational techniques could lead to further optimization of more specific and potent andrographolide analogues toward SARS-CoV-2 Mpro.

Evaluation of Apoptotic Activity of Ethanolic Extract of Andrographis paniculata Linn in Human Hepatocellular Carcinoma Cell Lines

Introduction: Andrographis paniculata is one of the highly used potential medicinal plants in the world. This plant is traditionally used for the treatment of common cold, diarrhoea, fever due to several infective cause, jaundice, as a health tonic for the liver and cardiovascular health, and as an antioxidant. It is also used to improve sexual dysfunctions and serve as a contraceptive. All parts of this plant are used to extract the active phytochemicals, but the compositions of phytoconstituents widely differ from one part to another and with place, season, and time of harvest.
 Materials and Methods: All data obtained were analysed by Student t test using MS- Excel. The results were computed statistically using one way ANOVA.The reagents used were MTT [3-(4,5-dimethythiazol-2-yl) 2,5-diphenyl tetrazolium bromide]: 0.5 mg MTT/ml of serum-free DMEM, Solubilization solution: Dimethyl sulfoxide (DMSO)and Phosphate Buffered Saline (PBS; pH 7.4).The cytotoxic effects of lycopene Ag NP on 3T3 cells. Cells were treated with Lycopene Ag NP(10, 20, 30, 50, 100 and 200 μM) for 24 h, and cell viability was evaluated by MTT assay. Data are shown as means ± SD (n = 3). * compared with the control-blank group, p < 0.001.
 Results: The results were evaluated using three tests namely, MTT assay, Dual staining (acridine orange/ethidium bromide) and morphological changes.
 Conclusion: Different stages of apoptosis revealed the importance of A.paniculata in hepatocellular carcinoma cell lines. Andrographolide and analogues have been subjected to extensive chemico- biological investigations for anticancer drug development. Several andrographolide analogues have shown superior anticancer activities in both in vitro and in vivo models. Further clinical and biomedical studies are required to confirm the pharmacological, pharmaceutical and toxicological properties of andrographolide.

The Andrographolide Analogue 3A.1 Synergizes with Taxane Derivatives in Aggressive Metastatic Prostate Cancers by Upregulation of Heat Shock Proteins and Downregulation of MAT2A-Mediated Cell Migration and Invasion.

Conventional treatment with taxanes (docetaxel-DTX or cabazitaxel-CBZ) increases the survival rates of patients with aggressive metastatic castration-resistant prostate cancer (mCRPC); however, most patients acquire resistance to taxanes. The andrographolide analog, 19-tert-butyldiphenylsilyl-8,7-epoxy andrographolide (3A.1), has shown anticancer activity against various cancers. In this study, we investigated the effect of 3A.1 alone and in combination with DTX/CBZ against mCRPC and their mechanism of action. Exposure to 3A.1 alone exhibited a dose- and time-dependent antitumor activity in mCRPC. Chou-Talalay's combination index (CI) values of all 3A.1 + TX combinations were less than 0.5, indicating synergism. Co-treatment of 3A.1 with TX reduced the required dose of DTX and CBZ (P < 0.05). Caspase assay (apoptosis) results concurred with in vitro cytotoxicity data. RNA sequencing (RNAseq), followed by ingenuity pathway analysis (IPA), identified that upregulation of heat-shock proteins (Hsp70, Hsp40, Hsp27, and Hsp90) and downregulation of MAT2A as the key player for 3A.1 response. Furthermore, the top treatment-induced differentially expressed genes (DEGs) belong to DNA damage, cell migration, hypoxia, autophagy (MMP1, MMP9, HIF-1α, Bag-3, H2AX, HMOX1, PSRC1), and cancer progression pathways. Most importantly, top downregulated DEG MAT2A has earlier been shown to be involved in cell migration and invasion. Furthermore, using in silico analysis on the Cancer Genome Atlas (TCGA) database, this study found that MAT2A and highly co-expressed (r > 0.7) genes, TRA2B and SF1, were associated with worse Gleason score and nodal metastasis status in prostate adenocarcinoma patients (PRAD-TCGA). Immunoblotting, comet, and migration assays corroborated these findings. These results suggest that 3A.1 may be useful in increasing the anticancer efficacy of taxanes to treat aggressive PCa. SIGNIFICANCE STATEMENT: The andrographolide analogue, 19-tert-butyldiphenylsilyl-8,7-epoxy andrographolide (3A.1), showed anticancer activity against metastatic castration-resistant and neuroendocrine variant prostate cancers (mCRPC/NEPC). Additionally, 3A.1 exhibited synergistic anticancer effect in combination with standard chemotherapy drugs docetaxel and cabazitaxel in mCRPC/NEPC. Post-treatment gene expression studies revealed that heat shock proteins (Hsp70, Hsp40, Hsp27, and Hsp90) and MAT2A are important in the mechanism of 3A.1 action and drug response. Furthermore, DNA damage, cell migration, hypoxia, and autophagy were crucial pathways for the anticancer activity of 3A.1.

Abstract 1355: A novel andrographolide analogue (3A.1) synergizes with Taxane derivatives in aggressive metastatic prostate cancers through upregulation of heatshock proteins and downregulation of MAT2A-mediated cell migration and invasion

Abstract BACKGROUND: Prostate cancer (PCa) is the 2nd leading cause of non-cutaneous cancer deaths among men in the USA with many progressing to aggressive metastatic castration resistant PCa (mCRPC; PCa unresponsive to androgen deprivation). Conventional treatment with taxanes (TX; docetaxel (DTX) or cabazitaxel (CBZ)) increases survival rates only slightly. The andrographolide analog, 19-tert-butyldiphenylsilyl-8,7-epoxy andrographolide (3A.1) has shown anticancer activity against various cancers. In this study, we investigated the effect of 3A.1 alone and in combination with DTX and CBZ against models of aggressive PCa. METHODS: Androgen receptor negative (AR-ve) mCRPC cell lines were treated with CBZ, DTX and 3A.1 as single-agent or combination over a broad concentration range, and in vitro cytotoxicity was determined. Chou-Talalay's combination index (CI) theorem was used to determine synergism and predicted dose reduction of taxanes. Post-treatment effect of single-agent and combination regimens on gene expression profile (GEP) was assessed using mRNA sequencing. Differentially expressed genes (DEGs) and molecular pathways involved in 3A.1 mechanism of action and drug synergy were identified using DESeq2, edgeR and Ingenuity pathway analysis (IPA). Protein expression of top DE genes was confirmed by immunoblotting. Cell cycle analysis, scratch/wound healing, and COMET assays were used to functionally validate the top treatment-associated genes. RESULTS: Exposure to 3A.1 alone exhibited a dose- and time-dependent antitumor activity in mCRPC. CI values of all 3A.1+ TX combinations were less than 0.5, indicating synergism. Co-treatment of 3A.1 with TX reduced the required dose of DTX by 9.5 to 18-folds (p&amp;lt;0.05), and CBZ by 3 to 14-folds (p&amp;lt;0.05). Our caspase 3/7 assay (apoptosis) results agreed with in vitro cytotoxicity data. RNAseq followed by IPA analysis identified that the top treatment-induced DEGs belong to cell migration, growth, apoptosis (MMP1, HSPB1, PSRC1), protein ubiquitination (HNRNPH2, HSPA1A/HSPA1B), and DNA repair (H2BC4, 11) pathways, along with cancer progression. Most importantly, our top downregulated DEG was MAT2A which has earlier been shown to be involved in cell migration and invasion. Our analysis using Immunoblotting, COMET, and Wound healing/Scratch (cell migration) assays corroborated with these findings. Finally, using in silico analysis on the TCGA (The Cancer Genome Atlas) database, we found that MAT2A and highly co-expressed (r&amp;gt;0.7) genes, TRA2B and SF1, were associated with worser Gleason score and nodal metastasis status in prostate adenocarcinoma patients (PRAD-TCGA). CONCLUSION: These results suggest that 3A.1 may be useful in increasing the anticancer efficacy of taxanes to treat aggressive prostate cancer. Citation Format: Taraswi Mitra Ghosh, Teeratas Kansom, Suman Mazumder, Joshua Davis, Ahmed Alnaim, Aedan Bird, P Opanasopit, Amit K. Mitra, Robert Arnold. A novel andrographolide analogue (3A.1) synergizes with Taxane derivatives in aggressive metastatic prostate cancers through upregulation of heatshock proteins and downregulation of MAT2A-mediated cell migration and invasion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1355.

Toxicological and pharmacokinetic analysis at therapeutic dose of SRS27, an investigational anti-asthma agent.

SRS27, an andrographolide analogue, had been proven to have therapeutic properties at a dose of 3mg/kg in both in vitro and in vivo asthma models of our previous study. The present study focuses on the pharmacokinetic and toxicity profile of this compound to provide further evidence for the development of this compound as an anti-asthma agent. A simple pharmacokinetic study was performed in female BALB/c mice to measure blood plasma concentration of the compound at therapeutic dose. At a single dose of 3mg/kg, SRS27 had a relatively short half-life but was able to achieve a concentration range of 13-19μM that is related to its in vitro bioactivities. With regard to toxicity profile, SRS27 appears to be safe, as no histopathological changes were observed in the liver, kidneys and ovaries of SRS27-treated female BALB/c mice. In addition, there was no significant change in the mean body weight and organ weight of the animals in the SRS27-treated groups compared with the vehicle-treated control group at the end of the treatment. This fully supports the absence of any significant changes in peripheral blood leukocyte counts of SRS27-treated mice. Rewardingly, this acute toxicity study also revealed that SRS27 has a wide therapeutic window as no toxicity symptoms were detected with a dose up to 60mg/kg daily when tested for 14days. These results provide strong justification for further investigation of SRS27 as a potential new anti-asthma agent.

Design and synthesis of C-12 dithiocarbamate andrographolide analogues as an anticancer agent

A series of 21 new analogues of C-12 dithiocarbamate andrographolide was designed and synthesized from natural andrographolide isolated from a common Thai plant, Andrographis paniculata. The reaction used to manipulate the andrographolide scaffold was conducted in one pot under mild reaction conditions. This avoided toxic catalysts and gave nearly quantitative yields of new analogues, generally without by-products and can be easily scaled -up for industrial processing. All new analogues were evaluated against nine cancer cell lines, some analogues exhibited greater selective cytotoxic activity to MCF-7 cancer cell than that of the parent andrographolide and cancer drugs.

Fabrication and characterization of andrographolide analogue (3A.1) nanosuspensions stabilized by amphiphilic chitosan derivatives for colorectal cancer therapy

Recently, 19-tert-butyldiphenylsilyl-8,17-epoxy andrographolide (3A.1) is an andrographolide analogue which was a promising chemotherapeutic agent against some types of cancer. However, this compound has an extremely low aqueous solubility and challenging problem that limits its biological activity and clinical application. In this study, 3A.1 nanosuspensions (3A.1-NS) were fabricated by an anti-solvent technique, using three amphiphilic chitosan copolymers as surface stabilizers: naphthyl-grafted succinyl chitosan (NSC), octyl-grafted succinyl chitosan (OSC), and benzyl-grafted succinyl chitosan (BSC). Subsequently, the 3A.1-NS were freeze-dried using 5% mannitol as a cryoprotectant to transform into powder. The physicochemical characteristics demonstrated that the 3A.1-NS with optimal drug to polymer ratio of 1.5:1 had spherical morphology with mean particle size in a nanoscale (<500 nm), narrow size distribution, high negative surface charge, and high drug content. The 3A.1-NS stabilized by NSC was well dispersed in aqueous medium and this formulation was physically and chemically stable at 4 °C for at least 6 months. The in vitro cytotoxicity result showed a significant increase in the anticancer activity of the 3A.1-NS stabilized by NSC derivatives against colorectal cancer (HCT116) cells. Based on these results, the 3A.1-NS stabilized by NSC could be used as a promising delivery system for colorectal cancer treatment.

Folate-Functionalized Amphiphilic Chitosan Polymeric Micelles Containing Andrographolide Analogue (3A.1) for Colorectal Cancer

A site-specific drug delivery system of anticancer agents has been delveloped to enhance the therapeutic efficacy and reduce toxicity to the normal tissue. Semi-synthetic andrographolide analogue 3A.1 (19-tert-butyldiphenylsilyl-8,17-epoxy andrographolide) is one of the potential natural anticancer compounds against many types of cancer including colorectal cancer cells. However, the clinical applications of this compound are limited because of low water solubility and lack of suitable delivery carriers. This study aimed to increase the aqueous solubility and improve the anticancer efficacy of 3A.1 via active targeting approaches. In this study, 3A.1 was loaded into the polymeric micelles self-assembled from N-naphthyl-N,O-succinyl chitosan (NSC). The micelles were conjugated with folate moiety (Fol-NSC) for targeting to the cancer cells. All of the 3A.1-loaded micelles were prepared by dropping method, and the physicochemical properties (size, charge, morphology, encapsulating efficiency, loading capacity), in vitro release behavior and in vitro anticancer activities against HT29 colorectal cancer cells were investigated. The 3A.1-loaded micelles were successfully formulated by dropping method using NSC or Fol-NSC. The micelles loaded with 40% initial 3A.1 showed the maximum encapsulating efficiency and loading capacity. The micelles were in the nanometer range, having a negative surface charge and a spherical structure. The colon site-specific release of the 3A.1 from the 3A.1-loaded micelles was obtained. The release of 3A.1 from the Fol-NSC micelles was slower than that from the NSC micelles. Moreover, the Fol-NSC micelles exhibited superior anticancer efficacy than that of the NSC micelles and free 3A.1. In conclusions, the 3A.1-loaded Fol-NSC micelles developed in the present study had suitable physicochemical properties. These nanocarriers may be a potential delivery system for targeted delivery of the 3A.1 to colorectal cancer cells.

Synthesis of andrographolide analogues and their neuroprotection and neurite outgrowth-promoting activities.

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease and remains incurable. Currently, neuronal injury and synapse loss have been considered to be main features in the pathophysiology of AD. Thus, modulation of neuronal survival and neurite outgrowth may represent an efficient strategy for the treatment of AD. Based on the isolates from the traditional medicine Andrographis paniculata, a series of andrographolide analogues were prepared and evaluated for the neuroprotection and neurotrophic activity. Two compounds (3 and 12) could effectively inhibit LPS-induced NO production and iNOS expression as well as proinflammatory cytokines TNF-α and IL-6. Moreover, pretreatment with 3 and 12 could protect neurons against microglia-mediated neurotoxicity. Further, H2O2- and 6-OHDA induced neurotoxicity in PC12 cells were also attenuated by the novel 12. Our next study indicated that compounds 1, 4 and 10 promoted NGF-induced neurite outgrowth in PC12 cells, with 10 the most potent. To clarify the underlying mechanisms of active compounds (3, 10 and 12), system pharmacology was employed. The results revealed that muscarinic acetylcholine receptors (mAChRs) may be the main targets of 12 against AD, while thyroid hormone signaling pathway was involved in the mechanisms of 10. These study point to the therapeutic potential of andrographolide analogues against AD.

Effect of Semi-synthetic Andrographolide Analogue-loaded Polymeric Micelles on HN22 Cell Migration

Semi-synthetic andrographolide (AG) analogue, namely 19-tert-butyldiphenylsilyl-8,17-epoxy andrographolide, or 3A.1, is an anticancer drug. However, the major problem of 3A.1 is poor water solubility hindering its clinical applications. To improve the water solubility and anticancer potency of this analogue, 3A.1-loaded polymeric micelles employing N-naphthyl-N-O-succinyl chitosan (NSCS) as amphiphilic copolymer were prepared by the dropping method. The morphology, particle size, entrapment efficiency (%EE), and loading capacity (%LC) were evaluated. The 3A.1-loaded NSCS micelles were successfully prepared. These micelles were nano-size (66.26 to 102.53 nm) and with a spherical shape, with negative surface charge (-30.50 to -22.23 mV). The 3A.1-loaded NSCS micelles with 40 % drug loading exhibited the maximum values of both %EE (90.84 %) and %LC (25.95 %), indicating that a high amount of 3A.1 could be entrapped into the NSCS micelles. In addition, in vitro anticancer activity and cell migration assay on HN22 cells were evaluated. The 3A.1-loaded NSCS micelles exhibited stronger anticancer effect and cell migration suppression than the free drug. Therefore, these NSCS micelles containing 3A.1 may be potential nanocarriers for the treatment of oral cancer.

Crystal structure and anti-inflammatory and anaphylactic effects of andrographlide sulphonate E in Xiyanping, a traditional Chinese medicine injection.

Andrographlide sulphonate E, namely sodium 9-dehydro-17-hydro-andrographolide-19-yl sulphate, was one of the major ingredients of Xiyanping injection. The present study aimed to demonstrate its suitability as a reference standard for use of quality control of this traditional Chinese medicine preparation made from andrographlide that has been widely used to treat various infectious diseases. The stable crystals were prepared for unambiguous elucidation of the chemical structure by comprehensive spectral and thermal analysis. The anti-inflammatory effects were investigated using in vitro and in vivo methods, and the potential allergenic risk related with safety was evaluated by in silico molecular docking analysis. The dihydrated sulphonate derivative could be present as orthorhombic crystals with stable three-dimensional supramolecular structure, providing it the favourable physico-chemical stability as reference substance. It exhibited potent anti-inflammatory activity both in vitro and in vivo, suggesting the potency responsible for clinic efficacy of Xiyanping. Molecular docking further demonstrated its low risk of allergic reaction, as well as the proposed mechanism of anaphylactic effect of andrographolide analogues. Dihydrated sodium 9-dehydro-17-hydro-andrographolide-19-yl sulphate may be the ideal reference standard for use in quality control of Xiyanping.

The anti-cancer activity of an andrographolide analogue functions through a GSK-3\u03b2-independent Wnt/\u03b2-catenin signaling pathway in colorectal cancer cells

The Wnt/β-catenin signaling pathway plays a key role in the progression of human colorectal cancers (CRCs) and is one of the leading targets of chemotherapy agents developed for CRC. The present study aimed to investigate the anti-cancer effects and molecular mechanisms of 19-O-triphenylmethyl andrographolide (RS-PP-050), an andrographolide analogue and determine its activity in the Wnt/β-catenin pathway. RS-PP-050 was found to potently inhibit the proliferation and survival of HT-29 CRC cells. It induces cell cycle arrest and promotes apoptotic cell death which was associated with the activation of PARP-1 and p53. Furthermore, RS-PP-050 exerts inhibitory effects on β-catenin transcription by suppressing T-cell factor/lymphocyte enhancer factor (TCF/LEF) activity in cells overexpressing β-catenin and by down-regulating the endogenous expression of Wnt target genes. RS-PP-050 also decreased the protein expression of the active form of β-catenin but functions independently of GSK-3β, a negative regulator of Wnt. Interestingly, RS-PP-050 extensively blocks phosphorylation at Ser675 of β-catenin which links to interference of the nuclear translocation of β-catenin and might contribute to Wnt inactivation. Collectively, our findings reveal the underlying anti-cancer mechanism of an andrographolide analogue and provide useful insight for exploiting a newly chemotherapeutic agent in Wnt/β-catenin-overexpressing CRC cells.

Anticancer Activity of A Silyl Andrographolide Analogue Mediated Through Wnt/β‐Catenin Signaling In Colon Cancer Cells

Hyperactivation of Wnt/β‐catenin signaling implicated in oncogenesis of colorectal cancer (CRC) is a potential molecular target for chemotherapy. An andrographolide analogue, 3A.1 (19‐tert‐butyldiphenylsilyl‐8, 17‐epoxy andrographolide) has previously been reported to be potently cytotoxic toward cancer cells by unknown molecular mechanisms. The present study explored the anti‐cancer activity of analogue 3A.1 on Wnt/β‐catenin signaling in colon cancer cells (HT29 cells) which were more sensitive to the others (HCT116 and SW480 cells). Analogue 3A.1 inhibited viability of HT29 cells with IC50 value of 11.1 ± 1.4 μM at 24 h, which was more potent than that of the parent andrographolide. Analogue 3A.1 also suppressed the proliferation of HT29 cells and induced cell apoptosis in a dose‐dependent manner. Its apoptotic activity was accompanied with increased expressions of proteins related to DNA damages; PARP‐1 and γ‐H2AX. In addition, analogue 3A.1 significantly inhibited T‐cell factor and lymphoid enhancer factor (TCF/LEF) promoter activity of Wnt/β‐catenin signaling. Accordingly, the expressions of Wnt target genes and β‐catenin protein were suppressed. Moreover, analogue 3A.1 increased the activity of GSK‐3β kinase, which is a negative regulator responsible for degradation of intracellular β‐catenin. This mode of action was further supported by the absence of the effects after treatment with a GSK‐3β inhibitor, and over‐expression of a mutant β‐catenin (S33Y). Our findings reveal, for the first time, an insight into the molecular mechanism of the anti‐cancer activity of analogue 3A.1 through the inhibition of Wnt/β‐catenin/GSK‐3β pathway and provide a therapeutic potential of the andrographolide analogue 3A.1 in CRC treatment.Support or Funding InformationThis research project was supported by TRF through the RGJ‐Ph.D. Program (PHD/0193/2553 to PP and SR) and the research grant IRN‐58W0004.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

A silyl andrographolide analogue suppresses Wnt/β-catenin signaling pathway in colon cancer

Hyperactivation of Wnt/β-catenin signaling implicated in oncogenesis of colorectal cancer (CRC) is a potential molecular target for chemotherapy. An andrographolide analogue, 3A.1 (19-tert-butyldiphenylsilyl-8, 17-epoxy andrographolide) has previously been reported to be potently cytotoxic toward cancer cells by unknown molecular mechanisms. The present study explored the anti-cancer activity of analogue 3A.1 on Wnt/β-catenin signaling in colon cancer cells (HT29 cells) which were more sensitive to the others (HCT116 and SW480 cells). Analogue 3A.1 inhibited viability of HT29 cells with IC50 value of 11.1 ± 1.4 μM at 24 h, which was more potent than that of the parent andrographolide. Analogue 3A.1 also suppressed the proliferation of HT29 cells and induced cell apoptosis in a dose-dependent manner. Its apoptotic activity was accompanied with increased expressions of proteins related to DNA damages; PARP-1 and γ-H2AX. In addition, analogue 3A.1 significantly inhibited T-cell factor and lymphoid enhancer factor (TCF/LEF) promoter activity of Wnt/β-catenin signaling. Accordingly, the expressions of Wnt target genes and β-catenin protein were suppressed. Moreover, analogue 3A.1 increased the activity of GSK-3β kinase, which is a negative regulator responsible for degradation of intracellular β-catenin. This mode of action was further supported by the absence of the effects after treatment with a GSK-3β inhibitor, and over-expression of a mutant β-catenin (S33Y). Our findings reveal, for the first time, an insight into the molecular mechanism of the anti-cancer activity of analogue 3A.1 through the inhibition of Wnt/β-catenin/GSK-3β pathway and provide a therapeutic potential of the andrographolide analogue 3A.1 in CRC treatment.

Review Literature: Andrographis paniculata

Andrographis paniculata (Burm f.) Nees is a medicinal plant used in many countries and used as a valuable traditional medicinal plant. It has many important bioactive compounds and is a potent drug used in Ayurveda, Siddha and Homoeopathy in many formulations. These medicinal plant is very effective in the treatment of various diseases like malaria, diabetes, viral hepatitis, cirrhosis, liver cancer, etc. The Andrographis paniculata (King of bitters), commonly known as Kalmegh is belongs to the family of acanthaceae and its major constituents are diterpenoids, flavonoids and polyphenols. Among the single compounds extracted from A. paniculata, and rographolide is the major one in terms of bioactive properties and the andrographolide analogues, impart bitter taste of this herb. The andrograp holides having constituents include 14-deoxy-12methoxyandrographolide, 14-deoxy-11, 12-didehydroandrographo; ides, 14-deoxyandrographolide. It cures and prevents a number of diseases in human beings and it is a boon of the nature of human healthy life. It prevents cold, fever, colic pain, active against inflammatory, antidiabetic activity, antioxidant, antifertility, cardiovascular and anti-virus including inhibited HIV. It is extensively used in indigenous system of medicines as home remedy for various diseases in Bangladeshi traditional system. It is used to treat hepatitis, gastrointestinal tract and upper respiratory infections, herpes, and a variety of other chronic and infectious diseases. The objective of this study is to review the literature of Andrographis paniculata specifically articles pertaining to therapeutic benefits, chemical properties and the attempt to provide recent update on phytochemical and pharmacological profile of Andrographis paniculata along its toxicity and condraindications status.

One-pot three steps cascade synthesis of novel isoandrographolide analogues and their cytotoxic activity

An efficient one-pot synthesis of novel andrographolide analogues is reported from a naturally occurring and abundant andrographolide isolated from aerial parts of Andrographis paniculata. Reactions in the one-pot proceed through a cascade epoxide ring opening by aniline derivatives/intramolecular ring closing and oxa-conjugate addition-elimination reactions. This methodology produces a new series of 17-amino-8-epi-isoandrographolide analogues in fair to excellent yields with high stereoselectivity using an economic and environmental procedure without base or catalyst at room temperature. Twenty-five analogues were obtained and cytotoxicity of all new analogues were evaluated against six cancer cell lines to search for a new lead compound based on andrographolide structure.

Inhibition of Topoisomerase IIα and Induction of Apoptosis in Gastric Cancer Cells by 19-Triisopropyl Andrographolide

Gastric cancer is the most common cancer in Eastern Asia. Increasing chemoresistance and general systemic toxicities have complicated the current chemotherapy leading to an urgent need of more effective agents. The present study reported a potent DNA topoisomerase IIα inhibitory activity of an andrographolide analogue (19-triisopropyl andrographolide, analogue-6) in gastric cancer cells; MKN-45, and AGS cells. The analogue was potently cytotoxic to both gastric cancer cell lines with the half maximal inhibitory concentration (IC50 values) of 6.3±0.7 μM, and 1.7±0.05 μM at 48 h for MKN-45, and AGS cells, respectively. It was more potent than the parent andrographolide and the clinically used, etoposide with the IC50 values of >50 μM in MKN-45 and 11.3±2.9 μM in AGS cells for andrographolide and 28.5±4.4 μM in MKN-45 and 4.08±0.5 μM in AGS cells for etoposide. Analogue-6 at 2 μM significantly inhibited DNA topoisomerase IIα enzyme in AGS cells, induced DNA damage, activated cleaved PARP-1, and Caspase3 leading to late cellular apoptosis. Interestingly, the expression of tumor suppressor p53 was not activated. These results show the importance of 19-triisopropyl-andrographolide in its emerging selectivity to primary target on topoisomerase IIα enzyme, inducing DNA damage and apoptosis by p53- independent mechanism. Thereby, the results provide insights of the potential of 19-triisopropyl andrographolide as an anticancer agent for gastric cancer. The chemical transformation of andrographolide is a promising strategy in drug discovery of a novel class of anticancer drugs from bioactive natural products.

The semi-synthesis of novel andrographolide analogues and anti-influenza virus activity evaluation of their derivatives.

Two novel andrographolide analogues with the structural motif of Δ(8,17)-alkene exo-to-endo isomerization, AI78 and AI89, were semi-synthesized firstly. Two series of derivatives were designed and synthesized based on the synthetic pathway (including series I: olefin isomerizing to endocyclic Δ(8,9) and series II: olefin isomerizing to endocyclic Δ(7,8)). The anti-influenza virus activity in vitro for all derivatives was evaluated. Among the compounds synthesized, compound 38 with benzyl amino group showed the greatest potency against H3N2 and was approximately 1.5-fold more potent than that of Lianbizhi, andrographolide analogue used clinically in China. Adamantyl derivative, 43, presented the lowest toxicity, with a higher TC50 and TI values than Lianbizhi. The structure-activity relationships studies of the synthetic analogues indicated that the endocyclic Δ(7,8)-double bond is preferable for anti-viral effect. Furthermore, the introduction of the fatty amino attached to the rigid skeleton at C-17 is beneficial for activity.