Digitoxin is a naturally occurring cardiac glycoside and anti-cancer agent extracted mianly from Digitalis lanata[1]. Digitoxin actively inhibits herpes simplex virus type 1 (HSV-1) replication with a EC50 value of 0.05M and exhibited the highest cytotoxicity against HeLa cervical cancer cells with an IC50 value of 28nM at 48h[2][3].
In vitro, human cervical cancer cell line HeLa cells were treated overnight with various concentrations of Digitoxin (0, 4, 20, or 100nM) for 24h or with 20nM Digitoxin for 12, 24, or 36h. Digitoxin caused DNA damage to block the cell cycle at the G2/M phase by triggering the activation of the ATM/ATR-CHK1/CHK2-Cdc25C signaling pathway[3]. Prostat cancer cells PC3 cells and HeLa cells were treated with digitoxin(10 and 25nM) for 24h. Fresh medium and drug were added every day. Cells were continuously treated in this manner for up to 3 days. Digitoxin inhibited NFκB-driven TGFBR2 expression, as well as Vimentin, while elevating E-cadherin expression. Digitoxin also significantly reduces HSPB1 mRNA and the HSPB1/RBFOX2 mRNA ratio in PC3 cells[4]. HeLa cervical cancer cells were treated with Digitoxin (25nM to 1μM; 24 h). Digitoxin caused dose-dependent cytotoxicity and activated intrinsic apoptosis via cleavage of caspase-9, caspase-3 and PARP. Carcinotoxic activity of Digitoxin includes suppression of NFAT-driven c-MYC expression[5].
In vivo, male and female Long Evans rats were injected i.p. once daily for 4 consecutive days with pregnenolone-16a-carbonitrile (PCN) (50mg/kg) then injected i.p. with an ethanolic solution of Digitoxin (1-10mg (1.3-13gmol)/kg) twenty-four hours after the last injection. PCN protected rats from Digitoxin toxicity, by elevating both liver microsomal cytochrome P-450p and UDP-GT-dt₁ activities[6]. Digitoxin (0, 3, 10 and 30μg per 100g of body weight) was administered to 3 cotton rats in one dose intraperitoneally 6.25h before intranasal infection with 107TCID50/100g of cotton rat with influenza strain A/Wuhan/H3N2/359/95 for 4 days. Digitoxin significantly and differentially suppressed levels of the cytokines TNFα, oncogene/keratinocyte chemoattractant (GRO/KC), MIP2, monocyte chemoattractant protein 1 (MCP1) and IFNγ, in the cotton rat lung[7].
References:
[1] Haux J. Digitoxin is a potential anticancer agent for several types of cancer. Med Hypotheses. 1999 Dec;53(6):543-8.
[2] Su C T, Hsu J T A, Hsieh H P, et al. Anti-HSV activity of digitoxin and its possible mechanisms. Antiviral Res. 2008 Jul;79(1):62-70.
[3] Gan H G, Qi M, Chan C P. et al. Digitoxin inhibits HeLa cell growth through the induction of G2/M cell cycle arrest and apoptosis in vitro and in vivo. Int J Oncol. 2020 Aug;57(2):562-573.
[4] Pollard B S, Suckow M A, Wolter W R, et al. Digitoxin Inhibits Epithelial-to-Mesenchymal-Transition in Hereditary Castration Resistant Prostate Cancer. Front Oncol. 2019 Aug 2:9:630.
[5] Yang Q F, Dalgard C L, Eidelman O, et al. Digitoxin induces apoptosis in cancer cells by inhibiting nuclear factor of activated T-cells-driven c-MYC expression. J Carcinog. 2013 May 20:12:8.
[6] Arlotto M P, Sonderfan A J, McKinney M M, Parkinson A. Digitoxin metabolism by liver microsomal cytochrome P-450 and UDP-glucuronosyltransferase and its role in the protection of rats from digitoxin toxicity by pregnenolone-16 alpha-carbonitrile. Arch Biochem Biophys. 1986 Nov 15;251(1):188-97.
[7] Pollard B S, Blancoi J C, Pollard J R. Classical Drug Digitoxin Inhibits Influenza Cytokine Storm, With Implications for Covid-19 Therapy. In Vivo. 2020 Nov-Dec;34(6):3723-3730.
Digitoxin是一种天然存在的心脏糖苷类和抗癌剂，主要从Digitalis lanata中提取[1]。Digitoxin能有效抑制1型单纯疱疹病毒（HSV-1）复制，其EC50值为0.05μM，并在48h内对HeLa宫颈癌细胞表现出最高的细胞毒性，IC50为28nM[2][3]。
体外实验中，人宫颈癌细胞系HeLa细胞分别用不同浓度Digitoxin（0、4、20或100nM）处理24h，或用20nM Digitoxin处理12、24或36h。Digitoxin通过激活ATM/ATR-CHK1/CHK2-Cdc25C信号通路造成DNA损伤，将细胞周期阻滞于G2/M期[3]。前列腺癌细胞PC3和HeLa细胞分别用Digitoxin（10和25nM）处理24h，每天更换新鲜培养基和药物，持续处理最长3天。Digitoxin抑制NFκB驱动的TGFBR2表达及Vimentin，同时上调E-cadherin表达，并显著降低PC3细胞中HSPB1 mRNA及HSPB1/RBFOX2 mRNA比值[4]。HeLa宫颈癌细胞用Digitoxin（25nM至1μM；24h）处理后，呈剂量依赖性细胞毒性，并通过裂解caspase-9、caspase-3与PARP激活内源性凋亡。Digitoxin的致癌毒性包括抑制NFAT驱动的c-MYC表达[5]。
体内实验中，雌雄Long Evans大鼠先连续4天每日腹腔注射孕烯醇酮-16α-碳腈（PCN）（50mg/kg），末次注射24h后腹腔注射Digitoxin乙醇溶液（1-10mg（1.3-13μmol）/kg）。PCN通过升高肝脏微粒体细胞色素P-450p和UDP-GT-dt₁活性，保护大鼠免受Digitoxin毒性[6]。Digitoxin（0、3、10和30μg/100g体重）以单次腹腔注射给予3只棉鼠，6.25h后以107TCID50/100g剂量经鼻感染A/Wuhan/H3N2/359/95流感毒株，持续4天。Digitoxin显著且差异性地抑制棉鼠肺内细胞因子TNFα、GRO/KC、MIP2、MCP1和IFNγ的水平[7]。