Natural Compounds from Herbs that can Potentially Execute as Autophagy Inducers for Cancer Therapy

Accumulated evidence indicates that autophagy is a response of cancer cells to various anti-cancer therapies. Autophagy is designated as programmed cell death type II, and is characterized by the formation of autophagic vacuoles in the cytoplasm. Numerous herbs, including Chinese herbs, have been applied to cancer treatments as complementary and alternative medicines, supplements, or nutraceuticals to dampen the side or adverse effects of chemotherapy drugs. Moreover, the tumor suppressive actions of herbs and natural products induced autophagy that may lead to cell senescence, increase apoptosis-independent cell death or complement apoptotic processes. Hereby, the underlying mechanisms of natural autophagy inducers are cautiously reviewed in this article. Additionally, three natural compounds—curcumin, 16-hydroxycleroda-3,13-dien-15,16-olide, and prodigiosin—are presented as candidates for autophagy inducers that can trigger cell death in a supplement or alternative medicine for cancer therapy. Despite recent advancements in therapeutic drugs or agents of natural products in several cancers, it warrants further investigation in preclinical and clinical studies.


Introduction
Cancer is a group of diseases involving out-of-control of cell growth due to the accumulation of defects, or mutations, in their DNA and with an impendence to invade or spread to other parts of the body [1]. In 2015, about 90.5 million people were diagnosed with cancer [2]. About 14.1 million new cases occur each year (not including skin cancer other than melanoma) [3]. Consequently, it causes about 8.8 million (15.7%) human deaths [4]. Anti-cancer drugs including 5-fluorouracil (5-FU), cisplatin, etoposide, paclitaxel, and doxorubicin are commonly used to treat various cancers, such as cisplatin and doxorubicin in ovarian cancer, 5-FU in colon and gastric cancer, paclitaxel and doxorubicin in breast cancer, and etoposide in small-cell lung cancer. However, these chemotherapeutic agents have evident side effects such as nausea, vomiting, loss of appetite, decreased immunity, oral ulcers, and other adverse effects [5]. In general, the anti-cancer drugs, such as cisplatin and doxorubicin favor abnormal triggering via programmed cell death (PCD) such as apoptosis, necrosis, necroptosis, and autophagy in normal cells as well as abolishing inflammation of damaged cells. Remarkably, apoptosis and

Natural Autophagy Inhibitor from Herbs
Currently, many herbs, including Chinese herbs, have been applied as cancer treatments to complement and act as alternative medicines, supplements, or nutraceuticals to dampen the aforementioned problems. It has been shown in the literature that many anti-cancer natural compounds and extracts could initiate autophagy in tumor cells. As summarized in this paper, tumor suppressive actions of natural product-induced autophagy may lead to cell senescence, provoke apoptosis-independent cell death, and complement apoptotic cell death by robust or target-specific mechanisms. Notably, natural compounds are fundamental for pharmacological treatments, and more than 50% of all anti-cancer drugs are of natural origins, or at least derived from scaffolds present in nature. Emerging research shows that molecules of natural origins are useful for preventive and therapeutic purposes by targeting essential hallmarks and enabling described characteristics. Moreover, natural compounds can change the differentiation status of selected cell types. In the past decade, some autophagy-related inducers and inhibitors have been extensively investigated. Of note, some autophagy inhibitors including 3-methyladenine (3-MA), pepstatin A, bafilomycin A1, and betulinic acid have been well studied, and have been applied as suppressors to examine the modality of autophagy formation in autophagy research [11]. Curiously, other natural autophagy inhibitors have also been elucidated as: (1) Matrine, a natural compound extract used in traditional Chinese medicine, can modulate the maturation process of lysosomal proteases in gastric cancer cell line (SGC-7901) cells [12]; (2) Elaiophylin promotes autophagosome accumulation but blocks autophagic flux by attenuating lysosomal cathepsin activity, resulting in the accumulation of SQSTM1/p62 in various human ovarian cancer cell lines [13]; (3) Oblongifolin C (OC), a natural small molecule compound extracted from Garcinia yunnanensis Hu, is a potent autophagic flux inhibitor. Exposure to OC results in an increased number of autophagosomes and impaired degradation of SQSTM1/p62 [14]; (4) p53 siRNA and epigallocatechin gallate (EGCG) dual therapy leads to the activation of pro-apoptotic genes, the inhibition of pro-survival autophagy and cell network formation [15]; (5) Frondoside A, a triterpenoid saponin with a sugar-steroid structure, is derived from the orange-footed sea cucumber, Cucumaria frondosa and inhibits pro-survival autophagy, a known mechanism of drug resistance in the human urothelial carcinoma cell lines and showed the synergistic activity with cisplatin and gemcitabine [16]; and (6) Rhizochalinin (Rhiz) from the marine sponge Rhizochalina incrustata, a novel sphingolipid-like marine compound is characterized by a unique combination of anti-cancer properties via one scenario of pro-survival autophagy inhibition in the human prostate cancer cells [17]. Thereby, natural products have also been demonstrated as autophagy inducers.

Natural Autophagy Inducers from Herbs
Conversely, the autophagy cascade may play a regulatory or major role resulting in cell death, particularly when natural products are employed. Apparently, more recent studies have elicited the benefits and molecular mechanisms triggered by natural active components for anti-cancer activity, particularly in the induction of autophagy. Sirolimus, also known as rapamycin, is isolated from the bacterium Streptomyces hygroscopicus, and may be the most famous natural autophagy inducer in autophagy research [18]. The extract of Emblica officinalis (Amla) inhibits ovarian cancer (OC) cell growth in vitro and in vivo, possibly via inhibition of angiogenesis and activation of autophagy in OC [19]. The natural compound lipoic acid (LA) inhibits O (6)-methylguanine-DNA methyltransferase (MGMT) and induces autophagy and subsequently LA enhances the cytotoxic effects of temozolomide in . The combination of gossypol and BRD4770 increased LC3-II levels and the autophagosome number in PANC-1 cells. The compound combination appears to act in a BNIP3 (B-cell lymphoma 2 19-kDa interacting protein)-dependent manner, suggesting that these compounds act together to induce autophagy-related cell death in pancreatic cancer cells [21]. The synthesized natural alkaloid berberine derivatives are able to induce autophagy for human colon carcinoma HCT-116 and SW613-B3 cell lines  Table 1 summarizes our collection of compound names, herb sources, and cancer types to address natural autophagic inducers for anti-cancer activity.
3.2. 16-Hydroxycleroda-3,13-dien-15,16-olide (HCD) Polyalthia longifolia var. pendula Linn is popularly known as ulta Ashok in India and is widely grown in gardens in tropical and subtropical Asia, such as the southern part of Taiwan, Pakistan, and Sri Lanka, as an evergreen ornamental tree. Many parts of P. longifolia var. pendula Linn tree are important in traditional Indian medicine [94], and encompass various biological functions, such as anti-inflammatory activity in neutrophils, cytotoxicity towards breast cancer cells, and hepatoma cancer cells [95]. The bark has shown to have medicinal value in the treatment of skin diseases, fever, hypertension, diabetes, and helminthiasis [96]. Recently, the chemical components of P. longifolia var. pendula such as diterpenes (clerodane and triterpenes) and aporphine alkaloids have been isolated. Diterpenoids in the hexane extract of P. longifolia seeds has exhibited significant anti-bacterial and anti-fungal activity [97]. Clerodane diterpenes can induce apoptosis in human leukemia HL-60 cells [98]. Moreover, 16-Hydroxycleroda-3,13-dien-15,16-olide (HCD) and its analogs extracted from the bark of P. longifolia have strong anti-inflammatory activity [99]. The enhanced expression of cyto-protective HO-1 factor and anti-inflammatory enzyme in microglia has been reported [100]. Hereby, the induction of apoptosis in leukemia K562 cells via a reduction in histone-modifying enzymes, PRC2-mediated gene silencing, the reactivation of downstream tumor suppressor gene expressions [101], via the PI3K/Akt pathway, and Aurora B resulting in gene silencing and cell cycle disturbance [102]. Our previous studies have demonstrated that HCD could cause apoptosis of two brain cancer cell lines, N18 and C6, via inhibition of focal adhesion kinase (FAK)-related signaling pathway and accordingly induced the autophagic cell death through ROS generation and p38/ERK1/2 signaling pathway activation [103,104]. In oral squamous cell carcinoma, HCD induced autophagy by activating AMPKα and inhibiting Akt, PI3K-ClassIII, and Beclin-1 activity [105].

Prodigiosin (PG)
Prodigiosin (PG, PubChem CID: 5377753) is an alkaloid and natural red pigment, which is a secondary metabolite of Serratia marcescens and also from actinomycete bacteria [106]. It is characterized by a common pyrrolyl pyrromethene skeleton [107,108]. The biological role of these pigments in the producer organisms remains unclear. Bacterial PGs and their synthetic derivatives have antimicrobial (bactericidal and bacteriostatic) [109][110][111][112], antimalarial [109,110,113], and antitumor [109,110,[114][115][116] activities. In addition, they have been shown to be effective apoptotic agents against various cancer cell lines [117], with multiple cellular targets including multi-drug resistant cells with little or no toxicity towards normal cell lines and induce apoptosis in T and B lymphocytes [118,119]. Recently, PG can induce apoptosis in various cancer cells with low toxicity on normal cells and PG-induced apoptosis may ascribe to Bcl-2 and survivin inhibition in colorectal cancer HT-29 cells [120]. Moreover, PG and its structural analogue (compound R) have induced the expression of p53 target genes accompanied by cell-cycle arrest and apoptosis in p53-deficient cancer cells [121]. A previous study has indicated that PG could be effective as a potential inhibitor compound against COX-2 protein, and can be applied as an anti-inflammatory drug [122]. In melanoma cells, PG activates the mitochondrial apoptotic pathway by disrupting an anti-apoptotic member of the BCL-2 family-MCL-1/BAK complexes by binding to the BH3 domain [123]. Additionally, PG exerts nearly identical cytotoxic effects on the resistant cells in comparison to their parental lines to reveal that this pro-apoptotic agent acts independently on the overexpression of multi-drug resistance transporters-MDR1, BCRP, or MRP [124]. Mechanistically, PG engages the IRE1-JNK and PERK-eIF2α branches of the unfolded protein response (UPR) signaling to up-regulate CHOP that, in turn, mediates BCL2 suppression to induce cell death in multiple human breast carcinoma cell lines [125].

Perspectives: Natural Autophagy Inducers Potentiate a New Era of Chemotherapeutic Drug Discovery
This review is valuable in terms of clarifying important directions for research on the major role of autophagy inducers resulting in cell death and the underlying mechanisms as seen with numerous natural products. Notably, natural products are an important resource in the discovery of lead compound for anti-cancer drug development, and study on the role of autophagy in the tumor suppressive effects of natural products continues to produce insights into and emerging from difficulties. It is noteworthy that technical variations in detecting autophagy might affect data quality, and study should focus on elaborating the role of natural inducers in deciding cell fate. In vivo study monitoring of natural autophagy inducers in cancer treatment is expected to be a critical effort for the future. Furthermore, the clinically relevant action of autophagy-inducing natural products should be emphasized in translational study.
Author Contributions: Yaw-Syan Fu, May-Jywan Tsai, Henrich Cheng, and Ching-Feng Weng contributed to the literature collection, review and present idea; Shian-Ren Lin and Ching-Feng Weng wrote the paper; all authors have read and approved this manuscript.

Conflicts of Interest:
The authors declare no conflict of interest.