Cellular senescence and cancer: Focusing on traditional Chinese medicine and natural products

Abstract Cancer is the principal cause of death and a dominant public health problem which seriously threatening human life. Among various ways to treat cancer, traditional Chinese medicine (TCM) and natural products have outstanding anti‐cancer effects with their unique advantages of high efficiency and minimal side effects. Cell senescence is a physiological process of cell growth stagnation triggered by stress, which is an important line of defence against tumour development. In recent years, active ingredients of TCM and natural products, as an interesting research hotspot, can induce cell senescence to suppress the occurrence and development of tumours, by inhibiting telomerase activity, triggering DNA damage, inducing SASP, and activating or inactivating oncogenes. In this paper, the recent research progress on the main compounds derived from TCM and natural products that play anti‐cancer roles by inducing cell senescence is systematically reviewed, aiming to provide a reference for the clinical treatment of pro‐senescent cancer.


| Malignancy and current treatment
The incidence of malignant tumours is rising worldwide. Cancer is regarded as the leading cause of death and the most key obstacle to improve life expectancy in every country of the world in the 21st century. 1 Tumour is a new organism formed by the proliferation of local tissue cells under the action of various tumorigenic factors. The development of cancer consists of a number of complex stages of initiation, progression and promotion, in which the stage of progression is invertible which seems to be the phase for the most appropriate drug intervention. Although there are different types therapies targeting cancers, such as surgical resection, radiotherapy, chemotherapy, immunotherapy, biotherapy therapy, molecular-targeted therapy and treatment of TCM, however, not every therapy achieves the expected optimal effect results. For instance, surgical resection can remove the tumour, the risk of advanced cancer is increased, and the survival rate is reduced. Clinical combination of first-line chemotherapy drugs and radiotherapy can prevent the progress of cancer to some extent, but they may cause serious toxic effects, affect the metabolism and proliferation of normal tissues, and reduce the life quality of patients.
Furthermore, the limitation of radiotherapy is that it can cause hair loss, memory damage and even lead to a second cancer. The function of organs, such as brain, muscle and bone, will decline in the late radiotherapy. In addition, secondary cancer and normal tissue damage resulting from chemotherapy also bring clinical problems for the cancer survivor. In the course of chemotherapy, most chemotherapy drugs can cause bone marrow suppression, resulting in immune deficiency or decline. Some chemotherapy drugs can also cause liver toxicity, kidney toxicity, cardiac toxicity and so on. Even the low sensitivity of some cancer cells means that chemotherapy has little or no effect on overall survival, which may have a significant impact on prognosis. For immunotherapy, it is important to avoid autoimmune diseases during tumour treatment. Patients who benefit from these treatments cannot be predicted due to unclear therapeutic targets and mechanisms. Biological therapy and molecular-targeted therapy usually have little effect on middle and advanced cancer patients in most cases, and the therapeutic effect is not satisfactory. Currently, both treatments are prone to tumour recurrence and metastasis.
So far, there is no widely accepted optimal treatment option for cancer treatment. Hence, it is necessary to find out new treatment options and more effective treatment methods to treat malignant tumours, especially specific targeted drug therapies. Data from the WHO report that the most of global population living in developing countries rely on traditional medicines extracted from plants for primary health care. 2 Recently, studies have shown that many TCM and monomer compounds, such as flavonoids, phenylpropanoids and alkaloids, have significant anti-tumour effects. [3][4][5] Nowadays, TCM has been used for cancer care and acts as an active function in healthcare system due to their low toxicity, strong specificity and high efficacy. As an important source of active natural products, TCM has the characteristics of multi-component and multi-target, which could not only restrict the tumorigenesis, but also reduce the side effects of radiotherapy and chemotherapy. In addition, it also shows unique advantages in the tumour prevention and treatment, especially in enhancing the host immune function and prolonging the survival of patients. 6 At present, in clinical practice, TCM is often used to assist the application of chemotherapy drugs in order to enhance the efficacy of chemotherapy drugs and improve the quality of life of patients.

| Cell senescence and anti-tumour
Cell senescence is an irreversible cell cycle arrest state in which senescent cells permanently lose their capacity to proliferate. Senescent cells are characterized by morphological and metabolic changes, chromatin remodelling, gene expression changes and the emergence of a pro-inflammatory phenotype known as senescence-associated secretion phenotype (SASP). Cell senescence plays an important role in the prevention of cardiovascular, kidney, liver and other diseases, as well as tissue regeneration and repair, anti-tumour and other aspects. 7 Currently, cell senescence identifies as innate barrier in the formation of tumour and induces senescent growth stagnation of tumour cells, which has been a novel idea in anti-tumour research.
Interestingly, previous studies have shown the role of cell senescence in promoting cell death against tumour. Cyclin-dependent kinase 4 and 6 (CDK4/6) is a pivotal protein for cell cycle regulation, especially in phase G1 the mutation of which is closely related to the occurrence of tumour. According to the report, in the mouse models of KRAS mutant pancreatic ductal adenocarcinoma(PDAC), the combination of trametinib (MEK inhibitor) and palbociclib (CDK4/6 inhibitor) can induce PDAC cell senescence. The senescence-inducing treatment produces a SASP that includes angiogenic factors that promote tumour angiogenesis, thereby improving the efficacy of drug delivery and cytotoxic gemcitabine chemotherapy. 8 Specific E3 ubiquitin ligase, MDM2, can bind with wild-type p53 protein and inactivate it, that is, MDM2-mediated degradation of p53. Nutlin-3a, an MDM2 antagonist, can inhibit the degradation of p53, which enables p53 to play its role in inducing the senescence of leukaemia cells. Interestingly, the Nutlin-3a induced senescence of cells is irreversible. 9 Therefore, to balance the function between cell vitality and cell proliferation, inducing cancer cell senescence is a promising therapeutic model.
In addition to inhibiting the vitality and metastasis of cancer cells, regulating autophagy and apoptosis, blocking the cell cycle, active ingredients of TCM and natural medicine can also play an anti-tumour role by promoting cell senescence. The senescence process induced by TCM and natural drugs is relatively slow compared with other anti-tumour strategies, whereas a welcome benefit of it is that it does not cause widespread damage to surrounding tissues or skin. Therefore, the active components of TCM or natural drugs can induce the tumour cells senescence and selectively eliminate senescent cells, which can reduce inflammation and enhance the function of the immune system, thus delaying the progress of age-related diseases, improving health and prolonging life. The elderly cells activate the proliferative mechanisms of various organs to fight against cell senescence together, so that the cancer cells in the body have the opportunity to develop into tumours. As a consequence, selective scavenging of senescent cells is of great significance to prevent the occurrence and development of tumours. The anti-tumour effect of TCM through this mode tends to be in the treatment of middle and advanced cancer, and the long-term effect of which is much better. The outstanding advantage of TCM is to restore the balance in the body. For this reason, the paper summarizes the research status in regulating cell senescence of anti-tumour effects through TCM and natural products, in order to provide reference for the development of advanced cancer therapies by combining TCM with modern cutting-edge technologies.

| REG UL ATI ON OF CELLUL AR S ENE SCEN CE BY AC TIVE COMP OUNDS DERIVE S FROM TCM
Senescence is usually triggered by damaging stimuli, and cellular senescence can be divided into replicative senescence and stressinduced premature senescence. 10 In this section, the detailed mechanisms of TCM regulating senescence in cancer cells will be described. Meanwhile, the main biological phenomena and related proteins are roughly summarized for reference ( Figure 1).

| Inhibition of telomerase activity induces cell senescence
Normal non-mutant cells after undergoing 50 divisions (called Hayflick limit) will stop dividing, which is known as replicative senescence. Replication senescence is generally considered as one of the anti-cancer mechanisms of eukaryotes, which is caused by the shortening of telomeres during replication. During each round of DNA replication, telomeres gradually shorten, eventually reaching a critical length so that it could prevent further replication and thus stops the cell from division. Shorter and capped telomeres induce a DNA damage response, thereby triggering senescence. Telomere length is maintained by telomerase, which is expressed in the large majority of cancers cells. 11,12 Telomerase activity is insufficient to maintain balance with the rapid rate of cell proliferation, resulting in telomere shortening and cell ageing. 13 Tumour cells tend to increase telomere length and promote ageing-related genes mutations by activating telomerase so as to live on forever. 14 As a consequence, pre-senescence and tumour cells death can be induced by inhibiting telomerase activity and preventing telomere prolongation. 11,14 It has become one of the most promising strategies for cancer therapy to inhibit tumour generation via suppressing telomerase activity to induce cell senescence ( Figure 2).

| Phenolics
Ginger (Zingiber officinale Roscoe), the fresh roots of the perennial herbs of the ginger family, has a long history of medicinal plant study. It also has been reported to exhibit various biological activities, such as antimicrobial, anti-tumour, anti-inflammatory activity and resistance to diabetes. 15, 16 Kaewtunjai et al found that subcytotoxic doses of Z officinale extract (ZOE) could cause telomere shortening and trigger cell senescence in A549 lung cancer cells.
Moreover, active compounds in ZOE (6-paradol, 6-shogaol) were determined, which may be responsible for the suppression of hTERT expression and telomerase activity. The result also revealed that ZOE had no acute toxicity in rats, and along with anticlastogenic effect against liver micronucleus formation. In conclusion, ZOE had been shown to inhibit telomerase activity and subsequently induce cell senescence, thereby demonstrating its remarkable anti-tumour property. 17 Pterostilbene (PT, a dimethyl ether analog of resveratrol) exhibits semblable pharmacological bioactivities. Compared with resveratrol, PT has better pharmacokinetic properties. 18 It has been indicated that an excellent interaction occurred between PT and the active site of telomerase. 19 Rong-Jane Chen et al 20 discovered that PT-induced senescence partly emerged via a p53-dependent mechanism as follows: PT → suppression of telomerase activity → DNA damage → ATM/ATR/p53 activation → p21 induction → blocked S phase → cellular senescence.

| Flavonoids
Baicalein, a phenolic flavonoid compound, which is an active ingredient of Scutellaria baicalensis Georgi, has been reported to possess various bioactivities 21,22 Baicalin, the conjugate of baicalein (baicalein-7-glucuronide), also exists in S baicalensis. 23 Cumulative evidence shows that both baicalin and baicalein exert powerful anti-cancer effects in various cancers. 24

| Terpenoids
Triptolide (TPL) is a typical diterpenoid isolated from Chinese medicinal herb Tripterygium wilfordii Hook F. 26 Researchers have suggested that TPL processes extensive activities including antitumour, anti-fertility and anti-cystogenesis effects. 27 The results have revealed that TPL could obviously increase p53 and p21 Kip1 , proving that TPL could block cell cycle. In short, TPL promoted cell senescence and inhibited the growth of liver cancer by inhibiting cyclin and activating AKT signalling pathway. In addition, TPL could suppress the expression of hTERT and telomerase activity, contributing to acceleration of cell senescence which resulted in inhibition of tumour growth. 28

| Alkaloids
Paclitaxel, as a natural diterpenoid, was exacted from the bark and trunk of Yew Pacific. Known as the best natural anti-cancer drug that has been discovered, it restrains depolymerization and improves the stability of tubulin ultimately contributes to inhibit the growth of tumour cells. 29,30 Multani et al studied and contrasted the effects of paclitaxel with its water-soluble complexes on chromosome morphology and apoptosis induction in K1735 clone X-21.
The results showed that paclitaxel and its water-soluble conjugates could inhibit the occurrence and development of tumour by inducing extensive telomere erosion. These compounds also could stimulate chromosomal fusion and telomerase dysfunction. 31 The main mechanism of telomerase activity regulates cell senescence in spite of interacting indirectly with telomeric sequences and dramatically reduce transcription level of hTERT, which is closely related to the inhibition of telomerase activity. On the other hand, they also found that prolonged treatment with low doses of papaverine could accelerate the senescence of HepG2cell and β-Galactosidase staining verified it. 35 These results well explained that papaverine could restrain the development of HCC by inhibiting telomerase activity and inducing cell senescence.
Harmine isolated from the seeds of Peganum harmala L. and Banisteriopsis caapi, which belongs to the natural β-carboline alkaloid. As a folk medicine, harmine shows various remarkable pharmacological effects, such as antimutagenic, antidepressant and antiplatelet. 36 Lei Zhao et al indicated that harmine arrested cell proliferation and induced cellular senescence in MCF-7 cells, confirmed by positive SA-β-gal staining result. In the research, according to the telomeric repeat amplication protocol assay, harmine induced a notable decrease of telomerase activity in MCF-7 cells, which was accompanied by a significant downregulation of all the hTERT subunits. 37 To sum up, harmine has a prominent effect on regulating cell senescence which is accompanied by a significant suppression of telomerase activity via overexpressed factor of p53/p21 pathway.

| TCM extracts
Tianshengyuan-1 (TSY-1) is exacted via a distillation process of multiple Chinese herbs, involving mangnolia officinalis, schisandra Chinensis, pericarpium citri reticulatae viride and almond. It has been reported that TSY-1, an agent used to treat bone marrow deficiencies, has an outstanding effect stimulating telomerase activity. 38,39 Weibo Yu et al discovered that TSY-1 suppressed telomerase activity in HL60 cells but increased telomerase activity in PBMCs and HSCs. The result confirmed that TSY-1 has a reverse effect on leukaemia cells with inherently high telomerase activities (telomerase inhibitory) and normal blood mononuclear and stem cells with low telomerase activities (telomerase activator). It seemed that the effect of TSY-1 on telomerase activity in different cell systems may be mediated by the methylation of the TERT promoter. Furthermore, the β-galactosidase reporter staining assay also revealed that the effect of TSY-1 on telomerase activity in connection with cell senescence. 40 Taken together, TSY-1 exhibited an effect on maintaining telomerase homoeostasis of telomerase activity for telomerasebased target treatment of bone marrow deficiency and cancer.
Telomerase is highly overexpressed in 85% of tumour but is almost undetectable in most normal tissues. Therefore, it is suitable to be an anti-cancer target. As mentioned above, a large number of researches have indicated that various native compounds could reduce telomerase activity and hTERT mRNA/protein levels. Another approach used to inhibit telomerase activity is to target telomere. In this case, inhibition of telomere extension is a potential anti-cancer therapeutic strategy. To some extent, we can develop telomerase inhibitors based on TCM. It should be noted that there are still many serious problems to be solved, such as which stage the telomerase activation will occur, whether anti-telomerase treatment will affect the immune response of the body, and when the active ingredients of TCM will inhibit telomerase activity so as to promote cell senescence and block tumour growth.

| Oncogene-induced Senescence
Senescence can also be triggered by the activation of oncogenes (oncogene-induced senescence (OIS)) and the loss of tumour suppressor gene (TSG), as a potent cell-independent anti-cancer mechanism. Senescence, which occurs in cells undergoing oncogenic signal transduction, is a response to prevent their transformation into malignant cells. It has been scientifically discovered that OIS is typically characterized by the upregulation of cell cycle inhibitors p16 INK4A and p21 CIP1 . 58 During the OIS period, cells undergo a variety of phenotypic changes. For example, chromatin recombination in senescent cells results in the formation of SAHF, which inhibits the growth of senescent cells by silencing proliferation-promoting genes. 59 OIS cells require the participation of the p19 ARF -p53 and p16-Rb tumour suppressor pathways, since the inactivation of these tumour suppressor pathways allows cells to bypass oncogenic Ras-induced senescence ( Figure 4). 60,61 Chansu, a traditional Chinese medicine prepared from the skin and auricular gland secretions of giant toads, has been used not only for treatment of various cardiac diseases, infection and pain for millennium but also for anti-tumour therapy. 62 According to previous researches, bufalin exerts anti-cancer effect by regulating cell differentiation, autophagy and cell cycle. 63 65 Their findings suggest that bufalin has potential anti-cancer effect by activating p53-senescence in prostate cancer patients. Baicalin extracted from Scutellaria root with outstanding biological activities. 66  Abundant evidences from molecular studies and animal models have shown the efficiency of AVN A for treatment of many diseases due to its anti-inflammatory, anti-oxidative and anti-cancer activities. 69 According to the enlarged cell size, increased number of blue cells stained with SA-β-gal and the positive staining with γH2AX, it was indicated that cell senescence occurred with treatment of AVN A.
Significant G1 phase arrest and increase of critical effectors of cellular senescence such as p27, p16 and p21 were observed after AVN A treatment. Moreover, AVN A treatment significantly increased the expression of miR-129-3p, which markedly repressed the E3 ubiquitin ligase Pirh2 and two other targets, IGF2BP3 and CDK6. The expression of p53 and p21 was increased after Pirh2 silencing by miR-129-3p, suggesting that the anti-tumour effect of AVN A is related to cell senescence. 70 The result indicate that AVN A could be a potential agent for colorectal cancer, attributing to the role of miR-129-3p/Pirh2/p53 axis in regulation of cellar senescence.

| Microenvironment and cellular senescence
One of the characteristics of senescent cells is that they remain metabolically activity to produce and secrete superfluous factors which can affect the tissue microenvironment in different modalities. 71  These researches clearly showed that the anti-tumour activities of TCM and the active components of natural products in the form of pro-senescence can be achieved by influencing the activity of telomerase, DNA damage and activation of oncogenes (Table 1).

| Other forms
Meanwhile, the mechanisms involved in these phenomena are summarized ( Figure 6).

F I G U R E 6
The mechanism of active components promoting the senescence resistance to cancer

CO N FLI C T O F I NTE R E S T
No potential conflicts of interest are disclosed.

AUTH O R CO NTR I B UTI O N S
HY contributed to conception and design. YL searched the literature.
YL, SY and HY wrote the manuscript. KW, JL, XB, RW, YQ, TW and HY critically viewed, edited and approved the manuscript. All authors read and approved the final manuscript.

DATA AVA I L A B I L I T Y S TAT E M E N T
Data sharing is not applicable to this article as no new data were created or analysed in this study.