Hydroxyzine Induces Cell Death in Triple-Negative Breast Cancer Cells via Mitochondrial Superoxide and Modulation of Jak2/STAT3 Signaling

Shakya Rajina,Park Gyu Hwan,Joo Sang Hoon,Shim Jung-Hyun,Choi Joon-Seok 한국응용약물학회 2022 Biomolecules & Therapeutics(구 응용약물학회지) Vol.30 No.6

Treatment of triple-negative breast cancer (TNBC) has been limited due to the lack of molecular targets. In this study, we evaluated the cytotoxicity of hydroxyzine, a histamine H1 receptor antagonist in human triple-negative breast cancer BT-20 and HCC- 70 cells. Hydroxyzine inhibited the growth of cells in dose- and time-dependent manners. The annexin V/propidium iodide double staining assay showed that hydroxyzine induced apoptosis. The hydroxyzine-induced apoptosis was accompanied down-regulation of cyclins and CDKs, as well as the generation of reactive oxygen species (ROS) without cell cycle arrest. The effect of hydroxyzine on the induction of ROS and apoptosis on TNBC cells was prevented by pre-treatment with ROS scavengers, N-acetyl cysteine or Mito-TEMPO, a mitochondria-targeted antioxidant, indicating that an increase in the generation of ROS mediated the apoptosis induced by hydroxyzine. Western blot analysis showed that hydroxyzine-induced apoptosis was through down-regulation of the phosphorylation of JAK2 and STAT3 by hydroxyzine treatment. In addition, hydroxyzine induced the phosphorylation of JNK and p38 MAPK. Our results indicate that hydroxyzine induced apoptosis via mitochondrial superoxide generation and the suppression of JAK2/STAT3 signaling.

Domperidone Exerts Antitumor Activity in Triple-Negative Breast Cancer Cells by Modulating Reactive Oxygen Species and JAK/STAT3 Signaling

Shakya Rajina,Byun Mi Ran,Joo Sang Hoon,Chun Kyung-Soo,Choi Joon-Seok 한국응용약물학회 2023 Biomolecules & Therapeutics(구 응용약물학회지) Vol.31 No.6

The lack of molecular targets hampers the treatment of triple-negative breast cancer (TNBC). In this study, we determined the cytotoxicity of domperidone, a dopamine D2 receptor (DRD2) antagonist in human TNBC BT-549 and CAL-51 cells. Domperidone inhibited cell growth in a dose- and time-dependent manner. The annexin V/propidium iodide staining showed that domperidone induced apoptosis. The domperidone-induced apoptosis was accompanied by the generation of mitochondrial superoxide and the down-regulation of cyclins and CDKs. The apoptotic effect of domperidone on TNBC cells was prevented by pre-treatment with Mito-TEMPO, a mitochondria-specific antioxidant. The prevention of apoptosis with Mito-TEMPO even at concentrations as low as 100 nM, implies that the generation of mitochondrial ROS mediated the domperidone-induced apoptosis. Immunoblot analysis showed that domperidone-induced apoptosis occurred through the down-regulation of the phosphorylation of JAK2 and STAT3. Moreover, domperidone downregulated the levels of D2-like dopamine receptors including DRD2, regardless of their mRNA levels. Our results support further development of DRD2 antagonists as potential therapeutic strategy treating TNBC.

장내세균에 의한 crocin 대사가 사염화탄소로 유발한 in vitro 지질과산화에 미치는 영향

라지나샤키아 ( Rajina Shakya ),정태천 ( Tae Cheon Jeong ) 한국동물실험대체법학회 2021 동물실험대체법학회지 Vol.15 No.1

Role of intestinal microbiota in drug metabolism has not widely been considered yet, despite of its importance in the biotransformation of xenobiotics. Crocin and its aglycone, crocetin, originally isolated from the fruit of Gardenia jasminoides, are used in treatment of inflammation, cancer, metabolic disorders, cardiovascular and neurological diseases. In this study, in vitro metabolism of crocin to crocetin by intestinal contents from antibiotic-treated rats were investigated by using HPLC. As consistent with the in vivo results reported previously, in vitro results also showed better metabolism of crocin to crocetin in intestinal contents isolated from normal rats than those from antibiotic-treated rats. Moreover, anti-oxidant effects of crocin and crocetin were compared to specify which compound would be pharmacologically more active in rat liver S-9 fractions that contain metabolic activation system. The results showed protective effects by both compounds against CCl4-induced depletion of glutathione and/or elevation of malondialdehyde. Overall, the results suggested that intestinal microbiota might be responsible for pharmacological actions of crocin, and that the present in vitro metabolic activation system would be a useful tool for studying the role of xenobiotic metabolism in pharmacology and toxicology, which can reduce the number of animals used.

A simple <i>in chemico</i> method for testing skin sensitizing potential of chemicals using small endogenous molecules

Nepal, Mahesh Raj,Shakya, Rajina,Kang, Mi Jeong,Jeong, Tae Cheon Elsevier 2018 Toxicology letters Vol.289 No.-

<P><B>Abstract</B></P> <P>Among many of the validated methods for testing skin sensitization, direct peptide reactivity assay (DPRA) employs no cells or animals. Although no immune cells are involved in this assay, it reliably predicts the skin sensitization potential of a chemical <I>in chemico</I>. Herein, a new method was developed using endogenous small-molecular-weight compounds, cysteamine and glutathione, rather than synthetic peptides, to differentiate skin sensitizers from non-sensitizers with an accuracy as high as DPRA. The percent depletion of cysteamine and glutathione by test chemicals was measured by an HPLC equipped with a PDA detector. To detect small-size molecules, such as cysteamine and glutathione, a derivatization by 4-(4-dimethylaminophenylazo) benzenesulfonyl chloride (DABS-Cl) was employed prior to the HPLC analysis. Following test method optimization, a cut-off criterion of 7.14% depletion was applied to differentiate skin sensitizers from non-sensitizers in combination of the ratio of 1:25 for cysteamine:test chemical with 1:50 for glutathione:test chemical for the best predictivity among various single or combination conditions. Although overlapping HPLC peaks could not be fully resolved for some test chemicals, high levels of sensitivity (100.0%), specificity (81.8%), and accuracy (93.3%) were obtained for 30 chemicals tested, which were comparable or better than those achieved with DPRA.</P> <P><B>Highlights</B></P> <P> <UL> <LI> An <I>in vitro</I> method to differentiate skin sensitizers and non-sensitizers is proposed by using cysteamine and glutathione. </LI> <LI> The study follows a simple covalent binding of a test chemical with low-molecular weight endogenous compounds. </LI> <LI> The proposed method presents sufficiently high sensitivity, specificity, and accuracy when compared with the existing methods. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Impact of gut microbiota on drug metabolism: an update for safe and effective use of drugs

노금한,강유라,Mahesh Raj Nepal,Rajina Shakya,강미정,강원구,이상규,정혜광,정태천 대한약학회 2017 Archives of Pharmacal Research Vol.40 No.12

The intestinal mucosa and liver have long beenconsidered as the main sites of drug metabolism, and thecontribution of gut microbiota to drug metabolism has beenunder-estimated. However, it is now generally acceptedthat the gut microbiota plays an important role in drugmetabolism prior to drug absorption or during enterohepaticcirculation via various microbial enzymatic reactionsin the intestine. Moreover, some drugs are metabolized bygut microbiota to specific metabolite(s) that cannot beformed in the liver. More importantly, the metabolism ofdrugs by gut microbiota prior to absorption can alter thesystemic bioavailability of certain drugs. Therefore,understanding drug metabolism by gut microbiota is criticalfor explaining changes in the pharmacokinetics ofdrugs, which may cause significant alterations in drug-inducedpharmacodynamics and toxicities. In this review, wedescribe recent progress with regard to the role of metabolismby gut microbiota in some drug-induced alterationsof either pharmacological or toxicological effects toemphasize the clinical importance of gut microbiota forsafe and effective use of drugs.

Phase I and phase II metabolite identification of rutaecarpine in freshly isolated hepatocytes from male Sprague–Dawley rats

이동욱,강유라,강미정,노금한,Mahesh R. Nepal,Rajina Shakya,이유정,장영동,이상규,김정애,정태천 대한약학회 2017 Archives of Pharmacal Research Vol.40 No.8

Rutaecarpine, an alkaloid originally isolatedfrom Evodia rutaecarpa, has been used for the treatment ofgastrointestinal disorders in Asia. In the present study, thephase I and phase II metabolites of rutaecarpine wereinvestigated in freshly isolated hepatocytes from maleSprague–Dawley rats. The individual metabolites werecharacterized via liquid chromatography-tandem massspectrometry. The incubation of rutaecarpine with freshlyisolated hepatocytes for 2 h yielded five major phase Imetabolites. In addition, three glucuronide conjugates andfour sulfate conjugates were observed. Because themajority of metabolites observed in vivo were identified,freshly isolated hepatocytes might be useful for the identificationof certain metabolites formed from drug candidatesfrom a reduced number of experimental animals.