Semi-synthetic Derivative Of Artemisinin Research Articles

Dihydroartemisinin suppresses the susceptibility of Anopheles stephensi to Plasmodium yoelii by activating the Toll signaling pathway

BackgroundMalaria is a serious public health concern. Artemisinin and its derivatives are first-line drugs for the treatment of Plasmodium falciparum malaria. In mammals, artemisinin exhibits potent anti-inflammatory and immunoregulatory properties. However, it is unclear whether artemisinin plays a regulatory role in the innate immunity of mosquitoes, thereby affecting the development of Plasmodium in Anopheles when artemisinin and its metabolites enter mosquitoes. This study aims to determine the effect of dihydroartemisinin (DHA), a first-generation semisynthetic derivative of artemisinin, on innate immunity and malaria vector competence of Anopheles stephensi.MethodsAnopheles stephensi was fed Plasmodium-infected mice treated with DHA via gavage, Plasmodium-infected blood containing DHA in vitro, or DHA-containing sugar, followed by Plasmodium yoelii infection. The engorged female mosquitoes were separated and dissected 8 and 17 days after infection. Plasmodium oocysts and sporozoites were counted and compared between the control and DHA-treated groups. Additionally, total RNA and proteins were extracted from engorged mosquitoes 24 and 72 h post infection (hpi). Real-time polymerase chain reaction (PCR) and western blotting were performed to detect the transcriptional levels and protein expression of immune molecules in mosquitoes. Finally, the Toll signaling pathway was inhibited via RNA interference and the infection density was analyzed to confirm the role of the Toll signaling pathway in the effect of DHA on the vector competence of mosquitoes.ResultsDHA treatment via different approaches significantly reduced the number of Plasmodium oocysts and sporozoites in mosquitoes. The transcriptional levels of anti-Plasmodium immune genes (including TEP1, LRIM1, and APL1C), Toll pathway genes (including Tube, MyD88, and Rel1), and the effector defensin 1 were upregulated by DHA treatment at 24 and 72 hpi. TEP1 and Rel1 protein expression was significantly induced under DHA treatment. However, Rel1 knockdown in DHA-treated mosquitoes abrogated DHA-mediated refractoriness to Plasmodium infection.ConclusionsDHA treatment effectively inhibited the development of P. yoelii in A. stephensi by upregulating mosquitoes’ Toll signaling pathway, thereby influencing the susceptibility of Anopheles to Plasmodium.Graphical

Dihydroartemisinin alleviates doxorubicin-induced cardiotoxicity and ferroptosis by activating Nrf2 and regulating autophagy.

Although the use of Doxorubicin (Dox) is extensive in the treatment of malignant tumor, the toxic effects of Dox on the heart can cause myocardial injury. Therefore, it is necessary to find an alternative drug to alleviate the Dox-induced cardiotoxicity. Dihydroartemisinin (DHA) is a semisynthetic derivative of artemisinin, which is an active ingredient of Artemisia annua. The study investigates the effects of DHA on doxorubicin-induced cardiotoxicity and ferroptosis, which are related to the activation of Nrf2 and the regulation of autophagy. Different concentrations of DHA were administered by gavage for 4 weeks in mice. H9c2 cells were pretreated with different concentrations of DHA for 24 h invitro. The mechanism of DHA treatment was explored through echocardiography, biochemical analysis, real-time quantitative PCR, western blotting analysis, ROS/DHE staining, immunohistochemistry, and immunofluorescence. Invivo, DHA markedly relieved Dox-induced cardiac dysfunction, attenuated oxidative stress, alleviated cardiomyocyte ferroptosis, activated Nrf2, promoted autophagy, and improved the function of lysosomes. Invitro, DHA attenuated oxidative stress and cardiomyocyte ferroptosis, activated Nrf2, promoted clearance of autophagosomes, and reduced lysosomal destruction. The changes of ferroptosis and Nrf2 depend on selective degradation of keap1 and recovery of lysosome. We found for the first time that DHA could protect the heart from the toxic effects of Dox-induced cardiotoxicity. In addition, DHA significantly alleviates Dox-induced ferroptosis through the clearance of autophagosomes, including the selective degradation of keap1 and the recovery of lysosomes.

Antitumour effects of artesunate via cell cycle checkpoint controls in human oesophageal squamous carcinoma cells

Artesunate (ART), an effective antimalarial semisynthetic derivative of artemisinin, exhibits antitumour properties, but the mechanism(s) involved remain elusive. In this study, we investigated the antitumour effects of ART on human oesophageal squamous cell carcinoma (ESCC) cell lines. Treatment of ESCC cell lines with ART resulted in the production of excessive reactive oxygen species (ROS) that induced DNA damage, reduced cell proliferation and inhibited clonogenicity via G1-S cell cycle arrest and/or apoptosis in vitro. The administration of ART to nude mice with ESCC cell xenografts inhibited tumour formation in vivo. However, the cytotoxicity of ART strongly differed among the ESCC cell lines tested. Transcriptomic profiling revealed that although the expression of large numbers of genes in ESCC cell lines was affected by ART treatment, these genes could be functionally clustered into pathways involved in regulating cell cycle progression, DNA metabolism and apoptosis. We revealed that p53 and Cdk4/6-p16-Rb cell cycle checkpoint controls were critical determinants required for mediating ART cytotoxicity in ESCC cell lines. Specifically, KYSE30 cells with p53Mut/p16Mut were the most sensitive to ART, KYSE150 and KYSE180 cells with p53Mut/p16Nor exhibited intermediate responses to ART, and Eca109 cells with p53Nor/p16Nor exhibited the most resistance to ATR. Consistently, perturbation of p53 expression using RNA interference (RNAi) and/or Cdk4/6 activity using the inhibitor palbociclib altered ART cytotoxicity in KYSE30 cells. Given that the p53 and Cdk4/6-cyclin D1-p16-Rb genes are commonly mutated in ESCC, our results potentially shed new light on neoadjuvant chemotherapy strategies for ESCC.

Dihydroartemisinin inhibits tumor progress via blocking ROR1-induced STAT3-activation in non-small cell lung cancer

In non-small cell lung cancer (NSCLC), identifying a component with certain molecular targets can aid research on cancer treatment. Dihydroartemisinin (DHA) is a semisynthetic derivative of artemisinin which induced the anti-cancer effects via the STAT3 signaling pathway, but the underlying molecular mechanism is still elusive. In this study, we first proved that DHA prohibits the growth of tumors both in vitro and in vivo. Data from transcriptomics showed that DHA reduced the expression level of the genes involved in cell cycle-promoting and anti-apoptosis, and most importantly, DHA restricted the expression level of receptor tyrosine kinase-like orphan receptor 1 (ROR1) which has been reported to have abnormal expression on tumor cells and had close interaction with STAT3 signaling. Then, we performed comprehensive experiments and found that DHA remarkably decreased the expression of ROR1 at both mRNA and protein levels and it also diminished the phosphorylation level of STAT3 in NSCLC cell lines. In addition, our data showed that exogenously introduced ROR1 could significantly enhance the phosphorylation of STAT3 while blocking ROR1 had the opposite effects indicating that ROR1 plays a critical role in promoting the activity of STAT3 signaling. Finally, we found that ROR1 overexpression could partially reverse the decreased activity of STAT3 induced by DHA which indicates that DHA-induced anti-growth signaling is conferred, at least in part, through blocking ROR1-mediated STAT3 activation. In summary, our study indicates that in NSCLC, ROR1 could be one of the critical molecular targets mediating DHA-induced STAT3 retardation.

Artemotil mediates protection in experimental colitis through the induction of Type 1 regulatory T cells

Abstract Artemotil (β-arteether) is a semi-synthetic derivative of artemisinin - a natural compound isolated from the plant Artemisia annuaand used worldwide as a front-line anti-malarial drug. Besides its anti-malarial effect, several studies have reported its potential use as an anti-tumor and anti-inflammatory agent. Artemotil was found to suppress the generation and function of Th1 and Th17 while promoting the generation of Foxp3+ regulatory T cells (Tregs). The role of Artemotil in modulating the generation and functions of Type 1 regulatory T cells is not yet identified. Tr1 cells are one of the key cell types that are essential for inhibiting inflammatory response through IL-10. In this study, we report an additional role of Artemotil by selectively inducing the generation of Tr1 cells induced by IL-27 by upregulating the signature of Tr1 cells genes such as c-maf and Prdm1, IRF-1, and Batf. We found that co-administer of Artemotil with anti-CD3 antibody increases the induction of IL-10 and frequency of Tr1 cells while suppressing Th1 and Th17 cells in vivo associated with less severe T-cell-induced enteropathy and intestinal inflammation. We further show the anti-inflammatory role of Artemotil in the chronic T-cell-induced colitis model in which Artemotil treatment alleviates the signs of colitis associated with the increased frequencies of Tr1 cells. Taken together, these data suggest that Artemotil provides protection in T-cell-mediated colitis by increasing the expansion of Tr1 cells and inhibiting the generation of Th1 and Th17 cells. This study was supported by grants from Science & Engineering Research Board (SERB), Department of Science and Technology (DST), Government of India (CRG/2018/002653)

Artesunate alleviates intracerebral haemorrhage secondary injury by inducing ferroptosis in M1-polarized microglia and suppressing inflammation through AMPK/mTORC1/GPX4 pathway.

Intracerebral haemorrhage (ICH) is a catastrophic subtype of stroke with severe morbidity and mortality. However, little progress has been made in the subsequent secondary injury. Artesunate, a water-soluble semi-synthetic derivative of artemisinin, exhibits remarkable pharmacological effects on anti-neuroinflammation. However, the effects of artesunate on ICH remain unknown. In the present study, haemoglobin (Hb) treatment in BV2 cell and collagenase type IV intracerebroventricular injection in Sprague-Dawley rats were used to establish in vitro and in vivo ICH models, respectively. For in vivo, the neurological scores, haematoma volume, brain oedema, inflammatory factors and iron deposition were evaluated. Besides, lipopolysaccharide (LPS) was used in in vitro to polarize BV2 cell to M1 phenotype. Cell viability, cellular reactive oxygen species (ROS), Fe2+ concentration, and lipid peroxidation levels, ferroptosis-associated proteins and mRNA, morphological of mitochondria were measured in vitro. Additionally, the AMP-activated protein kinase (AMPK)/mammalian/mechanistic target of rapamycin (mTOR) pathway were measured by western blot and immunofluorescence staining. The present in vivo results indicated that artesunate significantly ameliorated neurological deficits, haematoma volume and brain oedema in ICH rats. Besides, artesunate suppressed the M1-microglia relative inflammatory factors and up-regulated iron deposition. For in vitro, artesunate significantly selectively decreased the viability of LPS-stimulated BV2 cell. Furthermore, ROS and lipid peroxidation levels were up-regulated. And the glutathione peroxidase 4 (GPX4) were silenced via the AMPK/mTORC1 axis. Our finding supports that artesunate ameliorates the ICH secondary injury both in vitro and in vivo by inducing ferroptosis in microglia and further inhibiting inflammation mainly through the AMPK/mTORC1/GPX4 pathway. This finding may provide a novel target for ICH treatment.

TUG1/MAZ/FTH1 Axis Attenuates the Antiglioma Effect of Dihydroartemisinin by Inhibiting Ferroptosis.

Glioma is the most common primary intracranial malignant tumor in the brain. Currently, due to the limited treatment methods, the clinical outcome of patients with standard surgery combined with radiotherapy and chemotherapy is not satisfactory. Therefore, we urgently need to develop effective drugs to solve this problem. As a semisynthetic derivative of artemisinin, dihydroartemisinin (DHA) has been proved to have antitumor activity in glioma, which can induce apoptosis and inhibit the proliferation, migration, and invasion of glioma cells. In recent years, ferroptosis has been identified as another antitumor mechanism of DHA. Researchers have shown that DHA could promote ferroptosis in glioma cells. However, the specific molecular mechanisms of ferroptosis induced by DHA need more exploration. In this study, we found DHA could induce ferroptosis with ROS production and lipid peroxidation in glioma cells. Low expression of GPX4 and high expression of HMOX1 were identified in DHA treated glioma cells. Surprisingly, we found FTH1, a negative regulator of ferroptosis, upregulated in DHA treated glioma cells. It indicated that there should be some mechanisms that may cause ferroptosis attenuation in DHA treated glioma cells. For the first time, we confirmed that MYC-associated zinc finger protein (MAZ) could actively regulate FTH1 by binding to FTH1 promoter by CHIP assay. MAZ was further identified as the direct target of long noncoding RNA (lncRNA) TUG1 through luciferase assay. Downregulated expression of TUG1 and upregulated expression of MAZ were identified in DHA treated glioma cells. TUG1 overexpression or inhibition of FTH1 expression could enhance the antiglioma effect of DHA in vitro and in vivo, providing a promising strategy to enhance the antitumor effect of DHA in glioma.

LC-MS/TOF Characterization and Stability Study of Artesunate in Different Solvent Systems

Artemisinin (ART) is a sesquiterpene lactone and a popular malaria drug used in many parts of the world. Artesunate (ARTS) is a semi-synthetic derivative of ART with improved pharmacokinetic properties. However, the half-life of ARTS is less than an hour in vivo. The analysis of this drug in vitro in different solvent systems using LC-MS/TOF showed a solvent-driven breakdown. ARTS breakdown formed several derivatives, including dihydroartemisinin (DHA), artemether (ARTM) and DHA-dimer among others, at different rates in different solvent composition systems. The change in temperature from room temperature to physiological temperature (37 °C) was found to enhance the rate of the ARTS breakdown. In methanol, ARTS mainly formed ARTM with a chromatographic peak decrease of about 3.13%, while methanol and water (90:10) v/v mainly gave rise to DHA and ARTM with about an 80% chromatographic peak decrease. On the other hand, ARTS in methanol and ammonium acetate (85:15) v/v formed DHA, ARTM, DHA-dimer and other reaction peaks with about a 97% peak decrease and the formation of an orange solution pointing to a molecular re-arrangement reaction. These results have an important bearing on research on the analysis of artemisinin drugs conducted on these common solvents.

Dihydroartemisinin ameliorates chronic nonbacterial prostatitis and epithelial cellular inflammation by blocking the E2F7/HIF1α pathway.

Chronic nonbacterial prostatitis (CNP) has remained one of the most prevalent urological diseases, particularly in older men. Dihydroartemisinin (DHA) has been identified as a semi-synthetic derivative of artemisinin that exhibits broad protective effects. However, the role of DHA in inhibiting CNP inflammation and prostatic epithelial cell proliferation remains largely unknown. CNP animal model was induced by carrageenan in C57BL/6 mouse. Enzyme linked immunosorbent assay (ELISA), Real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot were used to examine inflammatory cytokines and proliferation genes expression. Immunofluorescence and immunochemistry staining were used to detect and E2F7 expression. Human prostatic epithelial cells (HPECs) and RWPE-1 was induced by lipopolysaccharide (LPS) to mimic CNP model in vitro. Cell proliferation was determined using MTS assay. DHA significantly alleviated the rough epithelium and inhibited multilamellar cell formation in the prostatic gland cavity and prostatic index induced by carrageenan. In addition, DHA decreased the expression of TNF-α and IL-6 inflammatory factors in prostatitis tissues and in LPS-induced epithelial cells. Upregulation of transcription factor E2F7, which expression was inhibited by DHA, was found in CNP tissues, human BPH tissues and LPS-induced epithelial cells inflammatory response. Mechanically, we found that depletion of E2F7 by shRNA inhibited epithelial cell proliferation and LPS-induced inflammation while DHA further enhance these effects. Furthermore, HIF1α was transcriptional regulated by E2F7 and involved in E2F7-inhibited CNP and cellular inflammatory response. Interestingly, we found that inhibition of HIF1α blocks E2F7-induced cell inflammatory response but does not obstruct E2F7-promoted cell growth. The results revealed that DHA inhibits the CNP and inflammation by blocking the E2F7/HIF1α pathway. Our findings provide new evidence for the mechanism of DHA and its key role in CNP, which may provide an alternative solution for the prevention and treatment of CNP.

Artesunate treatment ameliorates ultraviolet irradiation-driven skin photoaging via increasing β-catenin expression.

Objective: Artesunate, a semi-synthetic derivative of artemisinin, exerts various pharmacological activities. Nevertheless, the effects of Art on skin photoaging remain unclear. Herein, we investigated whether Art ameliorated ultraviolet-irradiated skin photoaging in HaCaT cells and mice.Methods: To construct skin photoaging cellular models, HaCaT cells were irradiated by UV (UVB, 20mJ/cm2) for 5 days. HaCaT cells were pretreated with three concentrations of Art (1, 5 and 20 μg/ml) for 2 h each day. After 5 days, cell senescence, ROS production, SOD levels, p16INK4a and β-catenin expression, proliferation and apoptosis were detected in HaCaT cells. Effects of Art on normal cells were investigated. After sh-β-catenin transfection or XAV-939 treatment, HaCaT cells were pretreated with 20 μg/ml Art and irradiated by UVB. After 5 days, skin photoaging was then observed. Furthermore, skin photoaging mouse models were established and the effects of Art and β-catenin silencing on skin photoaging were investigated.Results: Art treatment suppressed cell senescence, intracellular ROS production, p16INK4a expression and apoptosis and promoted proliferation and SOD and β-catenin expression in UVB irradiated HaCaT cells. But Art had no toxic effects on normal cells. Silencing β-catenin by sh-β-catenin or XAV-939 exacerbated UVB irradiation-mediated cell senescence, apoptosis, and ROS production in HaCaT cells, which was ameliorated by Art treatment. The therapeutic effects of Art on skin photoaging were also confirmed in mouse models.Conclusions: These findings suggested that Art treatment alleviated UVB irradiation-driven skin photoaging through enhancing β-catenin expression, which offered novel clues for pharmacological activity of Art.

Immunomolecular evaluation of dihydroartemisinin effects on apoptosis in chronic lymphocytic leukemia cell lines

IntroductionDihydroartemisinin (DHA), a semi-synthetic derivative of artemisinin, has recently shown to induce apoptosis in many types of cancer cells. In this study, we aimed to determine the effects of DHA on apoptosis in human chronic lymphocytic leukemia (CLL) cell lines. MethodsThe cells were treated separately and combined by DHA and Fludurabine (FLU) during 24, 48 and 72 hours. The cell viabilities determined by XTT method. Following separate and combined treatment of IC50 concentrations of DHA and FLU to the cells during 24 hours, the cells were analyzed by flow cytometry to determine the effects on apopotis staining with AnnexinV FITC and PI. mRNA and protein expression levels of TCTP, Mcl-1, Bcl-2, Bax and Caspase-3 were analyzed to find out the molecular mechanisms of apoptosis by using quantitative real-time PCR and flow cytometric methods. ResultsTreatment with DHA alone or in combination with FLU induced apoptosis in a dose dependent manner in CLL cells. DHA alone was more effective than FLU alone or combined treatment with DHA and FLU. Our results suggest that Bcl-2 protein family member Bax was active in the apoptotic response of CLL cells after DHA treatment. Moreover, the apoptotic response induced by DHA was independent from the p53 mutation status of the CLL cells. ConclusionDHA might be a potential anti-cancer therapeutic for CLL.

Enantioselective Synthesis and Profiling of Potent, Nonlinear Analogues of Antimalarial Tetraoxanes E209 and N205.

Synthetic endoperoxide antimalarials, such as 1,2,4-trioxolanes and 1,2,4,5-tetraoxanes, are promising successors for current front-line antimalarials, semisynthetic artemisinin derivatives. However, limited solubility of second-generation analogues in biological-relevant media represents a barrier in clinical development. We present methodology for the synthesis of nonlinear analogues of second-generation tetraoxane antimalarials E209 and N205 to investigate reduced molecular symmetry on in vitro antimalarial activity and physicochemical properties. While maintaining good antimalarial activity and metabolic stability, head-to-head comparison of linear and nonlinear counterparts showed up to 10-fold improvement in FaSSIF solubility for three of the four analogues studied. Pharmacokinetic studies in rats comparing a selected nonlinear analogue 14a and its parent N205 showed improvement on oral absorption and exposure in vivo with more than double the AUC and a significant increase in oral bioavailability (76% versus 41%). These findings provide support for further in vivo efficacy studies in preclinical animal species.

Artesunate inhibits melanoma progression in vitro via suppressing STAT3 signaling pathway.

Melanoma is a life-threatening cancer characterized with a potentially metastatic tumor of melanocytic origin. Improved methods or novel therapies are urgently needed to eliminate the development of metastases. Artesunate is a semi-synthetic derivative of artemisinin used for trarment of malaria and cancer. The purpose of this study was to investigate the anti-cancer effect of artesunate and the role on STAT3 signaling in A375 human melanoma cell line. Melanoma cells were treated with artesunate at concentrations of 0-5μM for 24 and 48h. The inhibition of cell viability, colony formation, migration, invasion, adhesion, percentage of apoptotic cells, and expressions of signal transducer and activator of transcription-3 (STAT3) and related proteins were examined. Artesunate inhibited cellular proliferation of cancer cells by induction of apoptosis at sub-toxic doses. Cells treated with artesunate showed an inhibition in adhesion to extracellular matrix substrate matrigel and type IV collagen. Artesunate treatment showed a decreased cellular migration, invasion, and colony formation in melanoma cells. Artesunate also inhibited STAT3 and Src activations and STAT3 related protein expressions; such as metalloproteinase 2 (MMP-2), MMP-9, Mcl-1, Bxl-xL, vascular endothelial growth factor (VEGF), and Twist. Moreover, overexpression of constitutively active STAT3 in A375 cells attenuated the anti-proliferative, apoptotic and anti-invasive effects of artesunate. The results obtained from this study demonstrated that the anticancer activity of artesunate occurred via STAT3 pathway and its target proteins. Therefore, it can be suggested that artesunate may be an important candidate molecule in the treatment of melanoma.

Dihydroartemisinin attenuated the symptoms of mice model of systemic lupus erythematosus by restoring the Treg/Th17 balance.

The Treg/Th17 imbalance is associated with the development of systemic lupus erythematosus (SLE). Dihydroartemisinin (DHA), a semi-synthetic derivative of artemisinin, is isolated from the traditional Chinese herb Artemisia annua Artemisia annua L. This study aims to evaluate the effects of DHA alone or in combination with prednisone in immunodeficiency of SLE. In vivo, the therapeutical effect of DHA alone or in combination with prednisone was assessed in the pristane-induced SLE mouse model. Then, the level of serum antibodies, creatinine (Cre), blood urea nitrogen (BUN), urine protein, kidney histopathology, interleukin (IL)-17, IL-6, transforming growth factor (TGF)-β, the expression of RORγt and Foxp3, the percentages of Treg and Th17 in peripheral blood and spleen were assayed. In vitro, the mouse spleen lymphocytes were separated and treated with DHA alone or DHA in combination with prednisone. Then the percentages of Treg and Th17, the concentration of IL-17, IL-6, TGF-β, and the expression of RORγt and Foxp3 were assayed. It was shown that DHA alone or in combination with prednisone treatment significantly alleviated the manifestations of pristane-induced SLE mice, suppressed inflammation and restored the Treg/Th17 balance. DHA alone or in combination with prednisone significantly inhibited Th17 cell differentiation while induced Treg cell differentiation in vitro. DHA alone or in combination with prednisone also reduced the transcription of RORγt and increased Foxp3 in lymphocytes, as well as IL-17 and TGF-β levels. Our data indicated that DHA can produce synergistic effect with prednisone to attenuate the symptoms of SLE by restoring Treg/Th17 balance.

Dihydroartemisinin inhibits the expression of von Willebrand factor by downregulation of transcription factor ERG in endothelial cells.

Dihydroartemisinin (DHA), a semi‐synthetic derivative of artemisinin, has effective antitumor and anti‐inflammatory actions. von Willebrand factor (vWF), a large multifunctional glycoprotein, has a prominent function in hemostasis and is a key factor in thrombus formation. In addition, vWF has been regarded as a prospective biomarker for the diagnosis of endothelial dysfunction. In our experiment, we observed that 25 μM DHA specifically downregulated the expression of vWF mRNA and protein in human umbilical vein endothelial cells (HUVECs). Further investigations demonstrated that this DHA‐decreased vWF expression was mediated by the transcription factor ERG and not GATA3. Luciferase activity assay confirmed that DHA regulated the ERG binding with the −56 ETS‐binding motif on the human vWF promoter. Thus, the −56 ETS motif on the vWF promoter region regulates the expression of vWF gene which is induced by DHA. Taken together, we proved that DHA decreased the vWF transcription through the downregulation of ERG in HUVECs. As vWF plays a key role in vascular homeostasis, our findings suggest a new role of DHA in vascular diseases.

Electrochemical and Theoretical Studies on the Artesunate Reduction

Malaria is the most devastating tropical disease in the world and this scenario is worsened by the absence of effective treatment. However, the plasmodium resistance to artemisinin does not show clinical relevance. The drug mechanism of action is associated to the endoperoxide moiety breakage. Artesunate is a semi-synthetic derivative of artemisinin and its absorption is facilitated due to its higher solubility in water. As a sesquiterpene lactone, artesunate can be electrochemically reduced in aqueous media on the glassy carbon electrode, having been studied by cyclic voltammetry, square wave voltammetry and chronoamperometry. The artesunate voltammetric reduction is diffusion-controlled and significantly irreversible. Its reduction is pH-independent, but from pH = 6.0 the cathodic current values increase in alkaline media, indicating that the proton-equilibrium occurs after the electron transfer step. The an values calculated vary from 0.30 to 0.37, leading to the 0.0975 s -1 value for k s . From the chronoamperometric data, two electrons (1.9 ± 0.4) are involved in the reduction process, being confirmed by the exact number of electrons obtained for artemisinin (1.9 ± 0.2). According to these results and computational findings, both drugs have the same reduction mechanism with the breakage of the endoperoxide bridge and consequent diol-derivative formation followed by the deoxy analog stabilization through the existence of a set of reactions involving protonation and charge transfer steps on the electrode surface. DOI: http://dx.doi.org/10.17807/orbital.v12i3.1439

Artesunate, as a HSP70 ATPase activity inhibitor, induces apoptosis in breast cancer cells

The present study aims to evaluate the inhibitory effects of artesunate (a semi-synthetic derivative of artemisinin) on HSP70 and Bcl-2 expression in two breast cancer cell lines, 4T1 and MCF-7. In addition, to determine in vitro inhibitory effect of artesunate against the ATPase activity of purified recombinant HSP70, it was tested in a carbonic anhydrase refolding assay with purified HSP70. Our results demonstrated that the artesunate not only induced apoptosis but also lead to the inhibition of HSP70 ATPase activity the in vitro (P < 0.001). The extent of HSP70 refolding inhibition increased with increasing μM concentrations of artesunate. Incubation of HSP70 with 50 μM artesunate showed significant inhibition of refolding activity by 38%. The IC50 values of artesunate for 4T1 cells, were lower than MCF-7 cells, indicating the higher sensitivity of the triple-negative phenotype. Furthermore, artesunate significantly down-regulated the expression of Bcl-2 and HSP70 while enhancing the expression of cleaved caspase-9 in MCF-7 and 4T1 cells. It also induced caspase-9 activity at 18 h in a dose-dependent manner in two breast cancer cell lines. Generally, our results show that the artesunate induces caspase-dependent apoptosis through the inhibition of HSP70 expression.

Dihydroartemisinin alleviates high glucose-induced vascular smooth muscle cells proliferation and inflammation by depressing the miR-376b-3p/KLF15 pathway

Inflammation and the proliferation of vascular smooth muscle cells (VSMCs) are seen to play critical roles in the development of vascular complications induced by diabetes and hyperglycemia. Dihydroartemisinin (DHA) has been identified as a semi-synthetic derivative of artemisinin that exhibits broad protective effects. However, the effect of DHA on high glucose (HG)-induced inflammation and proliferation of VSMCs remains unknown. Therefore, this study aims to show that DHA significantly inhibited the proliferation of VSMCs and that expression of the inflammatory cytokines IL-1β and TNF-α was induced by HG in a dose-dependent manner. Additionally, we were able to determine that KLF15 played a critical role in HG-induced VSMC proliferation and inflammation, confirming its protective effects observed after DHA treatment in the HG-induced inflammatory response of VSMCs. DHA was observed to directly depress the HG-induced expression of miR-376b-3p, which targeted the 3′-UTR of KLF15 and inhibited its expression. These results suggested that DHA plays a protective role in HG-induced VSMC proliferation and associated inflammation by inhibiting the miR-376b-3p/KLF15 axis. Our findings provide new evidence of the mechanisms of DHA and its critical role in treating the pathogenesis of diabetic vascular complications.

Artesunate induces apoptosis and inhibits the proliferation, stemness, and tumorigenesis of leukemia.

BackgroundLeukemia is characterized by the presence of highly malignant tumors formed in the hematopoietic system. Artesunate (Art), a semi-synthetic derivative of artemisinin, is commonly used as an antimalarial drug and has been proven to possess anticancer potential.MethodsIn this study, the effect of Art on the proliferation and stemness of human acute promyelocyte leukemia HL-60 cells and acute myeloid leukemia KG1a cells was investigated. Flow cytometry, colony formation assay, the protein expressive levels of survivin, P21, cleaved caspase 3, Bax, Bcl-2, Ki67 were detected the effect of Art on HL-60 and KG1a cells proliferation and apoptosis. At the same time, cell sphere formation assay and the protein expressive levels of CD44, SOX2, ALDH1 and OCT4 were used to analyze the effects of Art on cancer stem cell-like property in vitro. The orthotopic xenograft mouse models were established by using KG1a cells in BALB/c athymic nude mice. Tumor weigh was detected. The protein levels of survivin and Ki67 were detected by immunohistochemistry assays.ResultsArt induced cell apoptosis and inhibited cell proliferation and stemness in a dose-dependent manner. In the meantime, the results exhibited that Art inhibited the growth and stemness of transplanted tumors via the suppression of the MEK/ERK and PI3K/Akt pathway.ConclusionsOur present study provides new insights into the mechanisms of Art’s anticancer potential in leukemia.

Effects of dihydroartemisinin on the gut microbiome of mice.

Dihydroartemisinin (DHA) is a semisynthetic derivative of artemisinin, which has been found to exhibit a broad range of biological activities, excluding antimalarial effects; however its effects on the gut microbiota remain poorly understood. The present study aimed to investigate the effects of DHA on the gut microbiome in mice and to determine its potential biological and pharmaceutical activities through its alteration of the gut microbiota. Serum glucose, triglyceride (TG), total cholesterol, lipopolysaccharide, high density lipoprotein-cholesterol, low density lipoprotein-cholesterol, alanine aminotransferase and aspartate aminotransferase levels in mice treated with DHA were analyzed using the corresponding detection kits. In addition, hematoxylin and eosin staining was performed to determine the pathological effects of DHA on the liver, kidney and intestinal tissues of mice, and the effects of DHA on the gut microbiome were analyzed using 16S ribosomal (r)DNA gene analysis. The results demonstrated that the TG serum levels of mice treated with DHA were significantly decreased compared with the control group. Furthermore, 16S rDNA gene analysis demonstrated that the bacterial diversity of mice treated with DHA was enriched compared with the control group. The DHA group exhibited increased numbers of Firmicutes and Saccharibacteria, and decreased Deferribacteres and Actinobacteria compared with the control group at the phylum level. Kyoto Encyclopedia of Genes and Genomes signaling pathway enrichment analysis also revealed that the signaling pathways associated with ‘Energy metabolism’ and ‘Nucleotide metabolism’ were upregulated, whereas the signaling pathways associated with ‘Infectious diseases and ‘Neurodegenerative diseases’ were downregulated in the DHA group compared with the control group. In conclusion, the findings of the present study indicated that DHA may significantly decrease the serum TG levels and alter the gut microbiota, which suggested its potential to be used for the treatment of hyperlipidemia, inflammatory and neurodegenerative disorders.