Doxorubicin-induced senescence promotes stemness and tumorigenicity in EpCAM-/CD133- nonstem cell population in hepatocellular carcinoma cell line, HuH-7.

The therapeutic induction of senescence is a potential means to treat cancer, primarily acting through the induction of a persistent growth-arrested state in tumors. However, recent studies have indicated that therapy-induced senescence (TIS) in tumor cells allows for the prolonged survival of a subgroup of cells in a dormant state, with the potential to re-enter the cell cycle along with an increased stemness gene expression. Residual cells after TIS with increased cancer stem cell phenotype may have profound implications for tumor aggressiveness and disease recurrence. Herein, we investigated senescence-associated stemness in EpCAM+/CD133+ liver cancer stem cell and EpCAM-/CD133- nonstem cell populations in HuH7 cell line. We demonstrated that treatment with doxorubicin induces senescence in both cell populations, accompanied by a significant increase in the expression of reprogramming genes SOX2, KLF4, and c-MYC as well as liver stemness-related genes EpCAM, CK19, and ANXA3 and the multidrug resistance-related gene ABCG2. Moreover, doxorubicin treatment significantly increased EpCAM + population in nonstem cells indicating senescence-associated reprogramming of nonstem cell population. Also, Wnt/ß-catenin target genes were increased in these cells, while inhibition of this signaling pathway decreased stem cell gene expression. Importantly, Dox-treated EpCAM-/CD133- nonstem cells had increased in vivo tumor-forming ability. In addition, when SASP-CM from Dox-treated cells were applied onto hIPSC-derived hepatocytes, senescence was induced in hepatocytes along with an increased expression of TGF-ß, KLF4, and AXIN2. Importantly, SASP-CM was not able to induce senescence in Hep3B-TR cells, a derivative line rendered resistant to TGF-ß signaling. Furthermore, ELISA experiments revealed that the SASP-CM of Dox-treated cells contain inflammatory cytokines IL8 and IP10. In summary, our findings further emphasize the importance of carefully dissecting the beneficial and detrimental aspects of prosenescence therapy in HCC and support the potential use of senolytic drugs in HCC treatment in order to eliminate adverse effects of TIS.

Cellular Senescence Promotes Skin Carcinogenesis through p38MAPK and p44/42MAPK Signaling.

Cellular senescence entails an irreversible growth arrest that evolved in part to prevent cancer. Paradoxically, senescent cells secrete proinflammatory and growth-stimulatory molecules, termed the senescence-associated secretory phenotype (SASP), which is correlated with cancer cell proliferation in culture and xenograft models. However, at what tumor stage and how senescence and the SASP act on endogenous tumor growth in vivo is unknown. To understand the role of senescence in cancer etiology, we subjected p16-3MR transgenic mice, which permit the identification and selective elimination of senescent cells in vivo, to the well-established two-step protocol of squamous cell skin carcinoma, in which tumorigenesis is initiated by a carcinogen 7,12-dimethylbenz[α]anthracene, and then promoted by 12-O-tetradecanoyl-phorbol-13-acetate (TPA). We show that TPA promotes skin carcinogenesis by inducing senescence and a SASP. Systemic induction of senescence in nontumor-bearing p16-3MR mice using a chemotherapy followed by the two-step carcinogenesis protocol potentiated the conversion of benign papillomas to carcinomas by elevating p38MAPK and MAPK/ERK signaling. Ablation of senescent cells reduced p38MAPK and MAPK/ERK signaling, thereby preventing the progression of benign papillomas to carcinomas. Thus, we show for the first time that senescent cells are tumor promoters, not tumor initiators, and that they stimulate skin carcinogenesis by elevating p38MAPK and MAPK/ERK signaling. These findings pave the way for developing novel therapeutics against senescence-fueled cancers. SIGNIFICANCE: These findings identify chemotherapy-induced senescence as a culprit behind tumor promotion, suggesting that elimination of senescent cells after chemotherapy may reduce occurrence of second cancers decades later. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/17/3606/F1.large.jpg.