{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,27]],"date-time":"2026-01-27T23:25:20Z","timestamp":1769556320771,"version":"3.49.0"},"reference-count":54,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2021,10,1]],"date-time":"2021-10-01T00:00:00Z","timestamp":1633046400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100004663","name":"Ministry of Science and Technology, Taiwan","doi-asserted-by":"publisher","award":["MOST 107-2320-B-002-018-MY3"],"award-info":[{"award-number":["MOST 107-2320-B-002-018-MY3"]}],"id":[{"id":"10.13039\/501100004663","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Gongwin Biopharm Co., Ltd.","award":["2018ED0013"],"award-info":[{"award-number":["2018ED0013"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>Non-small cell lung cancer (NSCLC), an aggressive subtype of pulmonary carcinomas with high mortality, accounts for 85% of all lung cancers. Drug resistance and high recurrence rates impede the chemotherapeutic effect, making it urgent to develop new anti-NSCLC agents. Recently, we have demonstrated that para-toluenesulfonamide is a potential anti-tumor agent in human castration-resistant prostate cancer (CRPC) through inhibition of Akt\/mTOR\/p70S6 kinase pathway and lipid raft disruption. In the current study, we further addressed the critical role of cholesterol-enriched membrane microdomain and autophagic activation to para-toluenesulfonamide action in killing NSCLC. Similar in CRPC, para-toluenesulfonamide inhibited the Akt\/mTOR\/p70S6K pathway in NSCLC cell lines NCI-H460 and A549, leading to G1 arrest of the cell cycle and apoptosis. Para-toluenesulfonamide significantly decreased the cholesterol levels of plasma membrane. External cholesterol supplement rescued para-toluenesulfonamide-mediated effects. Para-toluenesulfonamide induced a profound increase of LC3-II protein expression and a significant decrease of p62 expression. Double staining of lysosomes and cellular cholesterol showed para-toluenesulfonamide-induced lysosomal transportation of cholesterol, which was validated using flow cytometric analysis of lysosome staining. Moreover, autophagy inhibitors could blunt para-toluenesulfonamide-induced effect, indicating autophagy induction. In conclusion, the data suggest that para-toluenesulfonamide is an effective anticancer agent against NSCLC through G1 checkpoint arrest and apoptotic cell death. The disturbance of membrane cholesterol levels and autophagic activation may play a crucial role to para-toluenesulfonamide action.<\/jats:p>","DOI":"10.3390\/molecules26195967","type":"journal-article","created":{"date-parts":[[2021,10,8]],"date-time":"2021-10-08T21:23:55Z","timestamp":1633728235000},"page":"5967","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["Autophagic Activation and Decrease of Plasma Membrane Cholesterol Contribute to Anticancer Activities in Non-Small Cell Lung Cancer"],"prefix":"10.3390","volume":"26","author":[{"given":"Jui-Ling","family":"Hsu","sequence":"first","affiliation":[{"name":"School of Pharmacy, National Taiwan University, No. 33, Linsen S. Rd., Zhongzheng Dist., Taipei 100, Taiwan"},{"name":"Department of Pharmacy, New Taipei Municipal TuCheng Hospital, Chang Gung Memorial Hospital, New Taipei City 236, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wohn-Jenn","family":"Leu","sequence":"additional","affiliation":[{"name":"School of Pharmacy, National Taiwan University, No. 33, Linsen S. Rd., Zhongzheng Dist., Taipei 100, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nan-Shan","family":"Zhong","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, Guangzhou Institute for Respiratory Health, First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510370, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6738-6054","authenticated-orcid":false,"given":"Jih-Hwa","family":"Guh","sequence":"additional","affiliation":[{"name":"School of Pharmacy, National Taiwan University, No. 33, Linsen S. Rd., Zhongzheng Dist., Taipei 100, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,10,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Testa, U., Castelli, G., and Pelosi, E. (2018). Lung cancers: Molecular characterization, clonal heterogeneity and evolution, and cancer stem cells. Cancers, 10.","DOI":"10.3390\/cancers10080248"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1007\/978-3-540-37696-5_2","article-title":"Genetic risk profiles for cancer susceptibility and therapy response","volume":"174","author":"Bartsch","year":"2007","journal-title":"Recent Results Cancer Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"669","DOI":"10.1016\/S1470-2045(05)70255-2","article-title":"Chemotherapy-induced neutropenia and treatment efficacy in advanced non-small-cell lung cancer: A pooled analysis of three randomized trials","volume":"6","author":"Gridelli","year":"2005","journal-title":"Lancet Oncol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e1800037","DOI":"10.1002\/ardp.201800037","article-title":"Overcoming resistance in non-small-cell lung cancer: A practical lesson for the medicinal chemist","volume":"351","author":"Ribaudo","year":"2018","journal-title":"Arch. Pharm."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"120858","DOI":"10.1172\/jci.insight.120858","article-title":"Personalized therapy for lung cancer: Striking a moving target","volume":"3","author":"Pakkala","year":"2018","journal-title":"JCI Insight"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"130","DOI":"10.1016\/j.jbior.2014.10.003","article-title":"Lipid rafts as major platforms for signaling regulation in cancer","volume":"57","author":"Mollinedo","year":"2015","journal-title":"Adv. Biol. Regul."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"491","DOI":"10.1093\/annonc\/mdr155","article-title":"Dietary cholesterol intake and cancer","volume":"23","author":"Hu","year":"2012","journal-title":"Ann. Oncol."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Ahmad, F., Sun, Q., Patel, D., and Stommel, J.M. (2019). Cholesterol metabolism: A potential therapeutic target in glioblastoma. Cancers, 11.","DOI":"10.3390\/cancers11020146"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2063","DOI":"10.1158\/0008-5472.CAN-15-2613","article-title":"The role of cholesterol in cancer","volume":"76","author":"Kuzu","year":"2016","journal-title":"Cancer Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"18777","DOI":"10.1074\/jbc.M611763200","article-title":"Squalene synthase, a determinant of Raft-associated cholesterol and modulator of cancer cell proliferation","volume":"282","author":"Brusselmans","year":"2007","journal-title":"J. Biol. Chem."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"675","DOI":"10.1164\/rccm.201404-0714OC","article-title":"Squalene synthase induces tumor necrosis factor receptor 1 enrichment in lipid rafts to promote lung cancer metastasis","volume":"190","author":"Yang","year":"2014","journal-title":"Am. J. Respir. Crit. Care Med."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1709","DOI":"10.1002\/jcp.22895","article-title":"Inhibition of lung cancer growth: ATP citrate lyase knockdown and statin treatment leads to dual blockade of mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K)\/AKT pathways","volume":"227","author":"Hanai","year":"2012","journal-title":"J. Cell Physiol."},{"key":"ref_13","first-page":"472","article-title":"Antitumor effect of para-toluenesulfonamide against lung cancer xenograft in a mouse model","volume":"5","author":"Gao","year":"2013","journal-title":"J. Thorac. Dis."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1026","DOI":"10.1097\/CAD.0000000000000283","article-title":"Para-toluenesulfonamide induces tongue squamous cell carcinoma cell death through disturbing lysosomal stability","volume":"26","author":"Liu","year":"2015","journal-title":"Anticancer Drugs"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1223","DOI":"10.3389\/fphar.2018.01223","article-title":"Para-toluenesulfonamide induces anti-tumor activity through Akt-dependent and \u2013independent mTOR\/p70S6K pathway: Roles of lipid raft and cholesterol contents","volume":"9","author":"Hsu","year":"2018","journal-title":"Front. Pharmacol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"6861","DOI":"10.3748\/wjg.v18.i46.6861","article-title":"Puncture injection of para-toluenesulfonamide combined with chemoembolization for advanced hepatocellular carcinoma","volume":"18","author":"He","year":"2012","journal-title":"World J. Gastroenterol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"838","DOI":"10.1097\/CAD.0b013e32832fe48f","article-title":"Gemcitabine plus cisplatin chemotherapy with concurrent para-toluenesulfonamide localinjection therapy for peripherally advanced nonsmall cell lung cancer larger than 3 cm in the greatest dimension","volume":"20","author":"He","year":"2009","journal-title":"Anticancer Drugs"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"2448","DOI":"10.21037\/jtd.2018.03.70","article-title":"Effects of para-toluenesulfonamide intratumoral injection on pulmonary adenoid cystic carcinoma complicating with severe central airway obstruction: A 5-year follow-up study","volume":"10","author":"Guan","year":"2018","journal-title":"J. Thorac. Dis."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1016\/j.lungcan.2016.05.012","article-title":"Effects of para-toluenesulfonamide intratumoral injection on non-small cell lung carcinoma with severe central airway obstruction: A multi-center, non-randomized, single-arm, open-label trial","volume":"98","author":"Li","year":"2016","journal-title":"Lung Cancer"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"781","DOI":"10.1080\/15384101.2016.1151581","article-title":"4E-BP1, a multifactor regulated multifunctional protein","volume":"15","author":"Qin","year":"2016","journal-title":"Cell Cycle"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"443","DOI":"10.3389\/fphar.2018.00443","article-title":"Inhibition of Akt\/mTOR\/p70S6K signaling activity with Huangkui capsule alleviates the early glomerular pathological changes in diabetic nephropathy","volume":"9","author":"Wu","year":"2018","journal-title":"Front. Pharmacol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"753","DOI":"10.1016\/j.bbagrm.2014.11.006","article-title":"Translational control by oncogenic signaling pathways","volume":"1849","author":"Gao","year":"2015","journal-title":"Biochim. Biophys. Acta."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"29","DOI":"10.2174\/157489210789702208","article-title":"Inhibitors of the PI3K\/Akt\/mTOR pathway: New hope for breast cancer patients","volume":"5","author":"Ghayad","year":"2010","journal-title":"Recent Pat. Anticancer Drug Discov."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1016\/S0005-2736(03)00016-6","article-title":"Role of cholesterol in lipid raft formation: Lessons from lipid model systems","volume":"1610","author":"Silvius","year":"2003","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_25","first-page":"2227","article-title":"Cholesterol-rich lipid rafts mediate akt-regulated survival in prostate cancer cells","volume":"62","author":"Zhuang","year":"2002","journal-title":"Cancer Res."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1136","DOI":"10.1038\/jid.2009.415","article-title":"Inhibition of Akt signaling by exclusion from lipid rafts in normal and transformed epidermal keratinocytes","volume":"130","author":"Calay","year":"2010","journal-title":"J. Investig. Dermatol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"246","DOI":"10.1016\/j.bbrc.2006.10.042","article-title":"Cholesterol depletion induces autophagy","volume":"351","author":"Cheng","year":"2006","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"249","DOI":"10.1016\/j.ejps.2009.07.010","article-title":"Involvement of PI3K-Akt-Bad pathway in apoptosis induced by 2,6-di-O-methyl-beta-cyclodextrin, not 2,6-di-O-methyl-alpha-cyclodextrin, through cholesterol depletion from lipid rafts on plasma membranes in cells","volume":"38","author":"Motoyama","year":"2009","journal-title":"Eur. J. Pharm. Sci."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"440","DOI":"10.1158\/0008-5472.CAN-09-1947","article-title":"Drug combination studies and their synergy quantification using the Chou-Talalay method","volume":"70","author":"Chou","year":"2010","journal-title":"Cancer Res."},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Lin, X., Liu, L., Fu, Y., Gao, J., He, Y., Wu, Y., and Lian, X. (2018). Dietary cholesterol intake and risk of lung cancer: A meta-analysis. Nutrients, 10.","DOI":"10.3390\/nu10020185"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Cancer Genome Atlas Research Network, Weinstein, J.N., Collisson, E.A., Mills, G.B., Shaw, K.R., Ozenberger, B.A., Ellrott, K., Shmulevich, I., Sander, C., and Stuart, J.M. (2013). The Cancer Genome Atlas Pan-Cancer analysis project. Nat. Genet., 45, 1113\u20131120.","DOI":"10.1038\/ng.2764"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"26","DOI":"10.1186\/s40364-018-0140-0","article-title":"Combination statin and chemotherapy inhibits proliferation and cytotoxicity of an aggressive natural killer cell leukemia","volume":"6","author":"Henslee","year":"2018","journal-title":"Biomark. Res."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"510","DOI":"10.4172\/1948-5956.1000316","article-title":"Targeting intracellular cholesterol is a novel therapeutic strategy for cancer treatment","volume":"6","author":"Mandal","year":"2014","journal-title":"J. Cancer Sci. Ther."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"471","DOI":"10.1016\/j.phrs.2018.11.035","article-title":"Cyclin-dependent protein serine\/threonine kinase inhibitors as anticancer drugs","volume":"139","author":"Roskoski","year":"2019","journal-title":"Pharmacol. Res."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Qiu, Z.X., Zhang, K., Qiu, X.S., Zhou, M., and Li, W.M. (2013). The prognostic value of phosphorylated AKT expression in non-small cell lung cancer: A meta-analysis. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0081451"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1016\/j.lungcan.2011.06.012","article-title":"Overexpression of the mammalian target of rapamycin (mTOR) and angioinvasion are poor prognostic factors in early stage NSCLC: A verification study","volume":"75","author":"Gately","year":"2012","journal-title":"Lung Cancer"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"3763","DOI":"10.1002\/cncr.25959","article-title":"Activation of mammalian target of rapamycin pathway confers adverse outcome in nonsmall cell lung carcinoma","volume":"117","author":"Liu","year":"2011","journal-title":"Cancer"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"34355","DOI":"10.1074\/jbc.M110.135103","article-title":"New hierarchical phosphorylation pathway of the translational repressor eIF4E-bindingprotein 1 (4E-BP1) in ischemia-reperfusion stress","volume":"285","author":"Ayuso","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"717","DOI":"10.1016\/j.molcel.2012.06.001","article-title":"Translational homeostasis via eIF4E and 4E-BP1","volume":"46","author":"Hinnebusch","year":"2012","journal-title":"Mol. Cell."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"833","DOI":"10.1203\/01.PDR.0000182588.20368.12","article-title":"Changes in 4E-BP1 and p70S6K phosphorylation in skeletal muscle of the ovine fetus after prolonged maternal fasting: Effects of insulin and IGF-I","volume":"58","author":"Shen","year":"2005","journal-title":"Pediatr. Res."},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Zeng, J., Zhang, H., Tan, Y., Sun, C., Liang, Y., Yu, J., and Zou, H. (2018). Aggregation of lipid rafts activates c-met and c-Src in non-small cell lung cancer cells. BMC Cancer, 18.","DOI":"10.1186\/s12885-018-4501-8"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"532","DOI":"10.1016\/j.bbamem.2013.07.018","article-title":"Interaction of membrane\/lipid rafts with the cytoskeleton: Impact on signaling and function: Membrane\/lipid rafts, mediators of cytoskeletal arrangement and cell signaling","volume":"1838","author":"Head","year":"2014","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"14509","DOI":"10.1073\/pnas.1019386108","article-title":"PI3K\/Akt signaling requires spatial compartmentalization in plasma membrane microdomains","volume":"108","author":"Gao","year":"2011","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"711","DOI":"10.1182\/blood-2006-04-016824","article-title":"Edelfosine and perifosine induce selective apoptosis in multiple myeloma by recruitment of death receptors and downstream signaling molecules into lipid rafts","volume":"109","author":"Gajate","year":"2007","journal-title":"Blood"},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Lima, R.T., Sousa, D., Gomes, A.S., Mendes, N., Matthiesen, R., Pedro, M., Marques, F., Pinto, M.M., Sousa, E., and Vasconcelos, M.H. (2018). The antitumor activity of a lead thioxanthone is associated with alterations in cholesterol localization. Molecules, 23.","DOI":"10.3390\/molecules23123301"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1038\/nrm3025","article-title":"mTOR: From growth signal integration to cancer, diabetes and ageing","volume":"12","author":"Zoncu","year":"2011","journal-title":"Nat. Rev. Mol. Cell. Biol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"81","DOI":"10.1038\/nrm3735","article-title":"Self-consumption: The interplay of autophagy and apoptosis","volume":"15","author":"Baehrecke","year":"2014","journal-title":"Nat. Rev. Mol. Cell. Biol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"7999","DOI":"10.1073\/pnas.1705304114","article-title":"mTORC1 activates SREBP-2 by suppressing cholesterol trafficking to lysosomes in mammalian cells","volume":"114","author":"Eid","year":"2017","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1038\/cr.2017.133","article-title":"Plasma membrane changes during programmed cell deaths","volume":"28","author":"Zhang","year":"2018","journal-title":"Cell Res."},{"key":"ref_50","doi-asserted-by":"crossref","unstructured":"Duarte, A., Poderoso, C., Cooke, M., Soria, G., Cornejo Maciel, F., Gottifredi, V., and Podest\u00e1, E.J. (2012). Mitochondrial fusion is essential for steroid biosynthesis. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0045829"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"463","DOI":"10.1002\/(SICI)1097-0029(19970315)36:6<463::AID-JEMT4>3.0.CO;2-J","article-title":"Roles of microfilaments and intermediate filaments in adrenal steroidogenesis","volume":"36","author":"Hall","year":"1997","journal-title":"Microsc. Res. Tech."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1007\/s10863-014-9592-6","article-title":"Mitochondrial cholesterol: Mechanisms of import and effects on mitochondrial function","volume":"48","author":"Martin","year":"2016","journal-title":"Bioenerg. Biomembr."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"5246","DOI":"10.1158\/0008-5472.CAN-07-6161","article-title":"Mitochondrial cholesterol contributes to chemotherapy resistance in hepatocellular carcinoma","volume":"68","author":"Montero","year":"2008","journal-title":"Cancer Res."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1200\/JCO.2004.08.001","article-title":"Gefitinib in combination with gemcitabine and cisplatin in advanced non-small-cell lung cancer: A phase III trial--INTACT 1","volume":"22","author":"Giaccone","year":"2004","journal-title":"J. Clin. Oncol."}],"container-title":["Molecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1420-3049\/26\/19\/5967\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:08:35Z","timestamp":1760166515000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1420-3049\/26\/19\/5967"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,10,1]]},"references-count":54,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2021,10]]}},"alternative-id":["molecules26195967"],"URL":"https:\/\/doi.org\/10.3390\/molecules26195967","relation":{},"ISSN":["1420-3049"],"issn-type":[{"value":"1420-3049","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,10,1]]}}}