GUO Mingyue, ZUO Ling, QIAO Gan, LIU Minghua, CAO Shousong,and LIN Xiukun, *

PI3K/Akt/mTOR Signaling as Targets for Developing Anticancer Agents from Marine Organisms

GUO Mingyue1), #, ZUO Ling1), #, QIAO Gan1), 2), LIU Minghua1), CAO Shousong1),and LIN Xiukun1), *

1),,,646000,2),646000,

The PI3K/Akt/mTOR signaling pathway is one of the most frequently dysregulated pathways in cancer. Targeting the PI3K-mediated pathway has been an important strategy for developing novel anticancer agents. In the past decades, more than 40 inhibitors of the PI3K/Akt/mTOR pathway have been developed at different clinical stages. Temsirolimus, everolimus, idelalisib, and copanlisib have been approved for clinical use by the Food and Drug Administration of the United States (FDA). However, the toxicity and drug resistance limit their efficiency in the treatment. Novel compounds with greater potency and selectivity, as well as improved therapeutic indices with reduced toxicity, are clearly required. Over the past three decades, a lot of bioactive ingredients with anticancer effects by affecting the PI3K-mediated pathways have been found from marine organisms. In the present mini-review, anticancer compounds from marine source that target the PI3K/Akt/mTOR signaling were reviewed. The molecular entities and their modes of action were presented. The marine compounds targeting special factors of the PI3K/Akt/mTOR were highlighted.

marine organisms; PI3K/Akt/mTOR pathway; anticancer activity

1 Introduction

The PI3K/Akt/mTOR pathway is one of the most frequently dysregulated pathways in cancer and, consequent- ly, more than 40 compounds that target key components of this signaling network have been tested in clinical trials involving patients with a range of different cancers (Janku, 2018). The clinical developments of many of these agents, however, have not advanced to late-phase rando- mized trials due to toxicity (Song, 2010). Presently, the mTOR inhibitors temsirolimus and everolimus and the PI3K inhibitors idelalisib and copanlisib have been app- roved by the FDA for clinical use in the treatment of a number of different cancers (Willis, 2020). Novel compounds with greater potency and selectivity, as well as improved therapeutic indices with reduced toxicity, are clearly required. In addition, biomarkers that are predictive of a response, such as PIK3CA mutations for inhibitors of the PI3K catalytic subunit α isoform, must be iden- tified and analytically and clinically validated (Wu, 2020). Finally, considering that oncogenic activation of the PI3K/Akt/mTOR pathway often occurs alongside pro-tu- morigenic aberrations in other signaling networks, rational combinations are also needed to optimize the effective-ness of treatment.

Marine organisms are rich sources for finding natural anticancer compounds. Due to their particular lifestyles in the sea with a special environment, some of the compounds such as bromophenols and depsipeptides are mostly found in marine sources. The biodiversity of marine organisms provides a rich source for the discovery and development of novel anticancer agents in the treatment of human malignancies (Zheng, 2018). Organisms living in the sea synthesize a wide variety of chemicals used as defense for predators (Wang, 2020). Compared with other biological components, some of the components frommarine organisms display more powerful cytotoxicity. Over the years, an increasing number of novel compounds with an- ticancer effects have been isolated from marine organisms, and many of them have been reported to possess promising anticancer activityinhibition of PI3K-mediated sig- naling. In this mini-review, we presented those compounds isolated from marine organisms with inhibitory effects on the PI3K/Akt/mTOR signaling. The challenges and prospects for developing anticancer agents using the PI3K/ Akt/mTOR signaling as target were also discussed.

2 Components Isolated from Marine Organisms Targeting the PI3K/-Akt/mTOR Pathway

As a natural carotenoid, fucoxanthin (Fig.1) is isolated from seaweed, and the compound exhibits a broad spectrum of biological activities, including anti-inflammatory (Heo, 2010), anti-mutagenic (Tanaka, 2012), and anticancer activities (Nishino, 1995; Satomi, 2017). Fu- coxanthin is capable of inhibiting the growth of a number of cancer cells, including liver cancer (Liu, 2009, 2013;Satomi and Nishino, 2009), colon cancer (Hosokawa, 2004; Liu, 2012), leukemia (Ishikawa, 2008; Yamamoto, 2011), prostate cancer (Satomi, 2012) and bladder cancer (Wang, 2012). It has been demonstrated that inactivation of the PI3K/Akt signaling is an important event in fucoxanthin-induced cancer cell apoptosis; treatment with fucoxanthin (1μmolL−1) led to down regulation of the p-PI3K, p-Akt significantly in HeLa can- cer cells (Ye, 2014). Additionally, exposure of hu- man glioma U251 and U87 cancer cells with fucoxanthin (50μmolL−1) resulted in an inhibitory effect of the Akt/ mTOR signaling and the compound also inhibited themigration and invasion of the cancer cells (Liu, 2016).ROS production is an important event in fucoxanthin- induced inhibitory effect on the invasion and migration of glioma cells; the inhibitory effect of the compound on can- cer cell growth was diminished in the presence of antioxidant glutathione (Wu, 2019). Recent study also found that fucoxanthin was capable of increasing the cytotoxicity of cisplatin; in the presence of fucoxanthin, the cell viability was markedly suppressed compared with that of cisplatin alone and the combination of fucoxanthin and cisplatin also increased the ratio of Bax/Bcl2 (Liu, 2013).

Fig.1 Chemical structures of compounds, isolated from marine organisms, targeting PI3K/Akt/mTOR signaling pathway.

Marine sponges are recognized as one of the most pro- ductive sources of anticancer components, and a lot of bio- active compounds with anticancer activity have been foundfrom marine sponges (Bader, 2005). Stellettin B (Stel B, Fig.1) belongs to an isomalabaricane triterpene. It has been extracted from marine spongePrevious study indicated that the compound was capable of inducing apoptosisthe PI3K/Akt/mTOR signaling pathway in cancer cells. Stel B treatment induced apoptosisenhancing the production of ROS, the cleavage of PARP, as well as inhibiting the activity of caspase 3/7 in human glioblastoma SF295 cells (Tang, 2014). Recent study showed that Stel B is able to inhibit the phosphorylation of PI3K and Akt in SF295 cells (1μmolL−1) and human chronic myeloid leukemia K562 cells (0.036μmolL−1) (Chen, 2017). Moreover, Stel B also leads to the ROS generation and increased expression of PARP in NSCLC cells (Hsiao, 2016). It can also induce G1 arrestinhibiting the expression of cyclin D1 and increasing the expression of p27 (Cheng, 2019). The PI3K/Akt/mTOR signaling pathway also plays a role in Stel B-induced (1μmolL−1) apoptosis in A549 cells. Additionally, Stel B treatment results in down-ex- pression of PI3K-p110 and inhibitory of the phosphorylation of PDK1, Akt, mTOR, and p70S6K (Hsiao, 2016).

In 1988, Rollisolated an alkaloid fascapysin from marine spongesp. (Roll, 1988). Re- cent study showed that its synthesized derivative 4-chloro fascaplysin (4-CF, 3μmolL−1, Fig.1) exhibited potent an- tiangiogenetic activityblocking the PI3K/Atk/mTOR pathway (Sharma, 2017). Treatment with 4-CF resulted in downregulation of VEGF and Akt in HUVEC cells, while in the presence of the inhibitors of VEGF and Akt (sunitinib and perifosine), the cytotoxic effect of 4- CF was significantly inhibited in HUVEC cells.

Sinulariolide (SNL, Fig.1), a cembrane-based diterpenoid, was isolated from cultured-type soft coral(Neoh, 2012). Previous studies have shown that SNL possesses anticancer effects by inducing apoptosis in several cancer cells; SNL treatment led to activation of caspase cascade in A375 melanoma cells (Li, 2013). Recent studies showed that the PI3K/Akt signaling played an important role in SNL-induced (10μgmL−1) cancer cell apoptosis in cancer cell invasion and migrationinhibiting the phosphorylation of PI3K, Akt and mTOR (Wu, 2015). In order to increase the efficiency of the compound, SNL was modified by conjugating with HA (hyaluronan) nanoparticles to form a novel complex compound HA/SNL aggregates. Since hyaluro- nan nanoparticles possesses high hydrophilicity, the bio- availability of the compound was greatly increased, and HA/SNL displays more powerful anticancer effect than SNL (Hsiao, 2016).

As an anticancer alkaloid, staurosporine (Fig.1) was ori- ginally isolated from a terrestrialsp. (Omu- ra, 1977). The compound was also isolated from several other marinesp. (Cartuche, 2019; Pimentel-Elardo, 2010). Staurosporine was capable of inducing apoptosis in human pancreatic carcinoma PaTu 8988t and Panc-1 cells. Staurosporine treatment resulted in decreased expression of Bcl-2 and Bad. The PI3K/Akt signaling plays an important role in stau- rosporine-induced cancer cell apoptosis. Treatment with staurosporine (20nmolL−1) can inhibite the Akt phosphoralytion significantly in HepG2 cells (Ding, 2017). To increase the anticancer activity, several derivatives of staurosporine were synthesized. Enzastaurin (Fig.1) is one of the derivatives. It can inhibit the cancer cell growth withpotent activity and higher water solubility (Faul, 2003). Enzastaurin (3.56μmolL−1) can inhibit the proliferation of gastric cancer bothandaffecting the Akt signaling cascade (Lee, 2008). Preclinical study showed that enzastaurin inhibited the growthof a panel of small-cell lung cancer lines. Enzastaurin treat- ment led to G1 arrest and apoptosis by suppressing the PKC/ERK 1/2 pathway in uveal melanoma cells carrying GNAQ mutation (Öztaşkın, 2015), and resulted in downregulation of the phosphorylation of GSK3βSer9, ri- bosomal protein S6Ser240/244, and AktThr308 significant- ly in xenogeneic glioblastoma cells. Enzastaurin also displays antiangiogenesis activity by downregulating VEGF (vascular endothelial growth factor) and suppressing the microvessel density in human tumor xenograft-bearing nude mice (Keyes, 2004). The safety of enzastaurin was confirmed by phase I clinical study for patients with refractory solid tumors and lymphoma. Oral administration of 500 mg of enzastaurin once daily were well tolerated (Li, 2016). Phase I clinical trial of the compound showed promise as minimal toxicity was demonstrated.However, phase II and phase III clinical trials is disappointing for patients with diffuse B cell lymphoma (Ro- bertson, 2007), relapsed and refractory mantle cell NSCLC (Oh, 2008) and lymphoma (Morschhauser, 2008). Therapy of enzastaurin with other chemotherapeutic agents were developed; the combination of en- zastaurin with bevacizumab for the treatment of recurrent malignant gliomas was well-tolerated, with similar results to bevacizumab monotherapy (Odia, 2016). Synergistic effects were also found using the combination of en- zastaurin and Ibrutinib in the treatment of large B cell lymphoma (He, 2019).

Bromophenols, a large class of marine natural products, are widely distributed in various marine organisms inclu- ding marine sponges, algae, cyanobacteria and marine fun- gi (Wang, 2013; Blunt, 2018). Bromophenols exhibits broad biological activities such as antibacterial activities, antivirus and anticancer activity (Liu, 2011; Öztaşkın, 2015; Wang, 2015). Shi’s laboratory developed a series of bromophenol derivatives and one of the bromophenol derivatives BOS-102 (Fig.1) displayed potent anticancer effect toward several human cancer cell lines especially for human lung cancer cells. Treatment with BOS-102 resulted in apoptosis and cell cycle arrest in lung cancer A549 cells. BOS-102 was able to activate caspase- 3 and PARP, enhance the Bax/Bcl-2 ratio, increase the ROS generation, and decrease ΔΨm, leading cytochrome release from mitochondria. Recent study revealed that the PI3K/Akt pathway was involved in the cancer cell apoptosis induced by BOS-102 (10μmolL−1). Treatment with the compound resulted in downregulation of the phospho- rylation of PI3K and Akt significantly in A549 cells (Guo, 2018).

Bostrycin (Fig.1) found in marine fungisp. in the South China Sea is able to inhibit the growth ofprostate and gastric cancer (Yang,2012; Fan, 2018). Studies have shown that bostrycin (10μmolL−1)can suppress the proliferation of human lung carcinoma A549 cellsdownregulation of the PI3K/Akt signaling. Treatment with bostrycin downregulated the levels of p- Akt significantly, while the levels of p27 were up-regu- lated, leading to cell cycle arrest at G1 phase (Chen, 2011).

As a sulfated polysaccharide compound, fucoidan (Fig.1) was originally found in the cell wall matrix of many kinds of brown seaweeds (Li, 2008). Fucoidan possesses a broad spectrum of bioactivities including antivirus, anti- bacterial and anti-cancer activities. In recent years, in- creasing attention is focused on the anti-cancer effect of fucoidan since the compound exhibits potent anticancer effect with low toxicity (Lee, 2004; Aisa, 2005; Gideon and Rengasamy, 2008; Yamasaki-Miyamoto, 2009; Yang, 2013). The PI3K/Akt signaling is invo- lved in fucoidan-induced cancer cell apoptosis in a num- ber of cancer cells. Treatment with fucoidan (200μgmL−1) suppressed the cell migration and viability significantly in HT-29 cells (Han, 2015). Fucoidan can also inhibit the growth of human breast cancer MDA-MB-231 cells by downregulating the expressions of p-PI3K, p-Akt, and p-GSK3β (Xue, 2017). Recent study also found that fucoidan was capable of suppressing the cell viability and inducing the apoptosis in human prostate PC-3 cancer cellsinhibiting the PI3K/Akt signaling pathway (Boo, 2013). Fucoidan-induced cancer cell apoptosis and cell cy- cle arrest are not associated with p53 expression. Treatments of the p53 positive and negative HCT-116 cells with fucoidan result in similar apoptosis and DNA damage (Park, 2017).

Xyloketal B (Fig.1) is obtained from mangrove fungussp. in the South China Sea (Lin, 2001). The compound exhibits antioxidant activity and protective ef- fects on endothelial and neuronal oxidative injuries. Xyloketal B (300μmolL−1) is capable of inhibiting the growth of glioma cells. The PI3K/Akt signaling is involved in the regulation of proliferation and migration of glioblastoma cells (Yajima, 2012). Treatment with xyloketal B leads to the downregulation of p-Akt and p-ERK1/2 sig-nificantly in human glioma U251 cancer cells (Chen, 2015).

3 Discussion

In the present mini-review, we discussed the compounds isolated from marine organisms and their main targets arethe PI3K/Akt/mTOR signalling (Table 1). However, it should be emphasized that some of the compounds can target multiple pathways. Except for targeting the PI3K/ Akt signaling, fucoxanthin can also suppress the p38 MAPK and NF-κB pathways; treatment with fucoxanthin can in- hibit the phosphorylation of MAPKs, and increase the expression of NF-κB-regulated Bax/Bcl-2 ratio (Kumar, 2013). ROS-mediated oxidative damage plays arole in fucoxian-induced cancer cell apoptosis (Jang, 2018). The PI3K/Akt/mTOR pathway is also involved in inflammatory event. The fucoxanthin can decrease pro-in- flammatory cytokines, such as IL-1b, IL-6, and TNF-α, by suppressing NO production and iNOS expression (Heo, 2010). Moreover, fucoxanthin can significantly in- hibit the inflammation of mice with paw edema by suppressing the levels of NO and Akt in the plasma. It can protect catalase (CAT) and superoxide dismutase (SOD) against disruption in mice with paw edema (Choi, 2016). More studies are needed to address if there is correlation between the anticancer activity and anti-inflam- matory effects of fucoxanthin.

Table 1 The compounds isolated from marine organisms and their main targets on the PI3K/Akt/mTOR signaling pathway

In the past decades, significant progress has been made in developing anticancer agents targeting the PI3K signalling. However, the toxicity and the drug resistance hinder the clinical application of these drugs. Several approaches overcoming the drug resistance are developing. It is important to find novel agents targeting alternative binding sites on the signalling factors or targeting the other pathways that are required for activating the PI3K signaling (Gumireddy, 2005). Future studies should focus on finding novel compounds from marine organisms with low toxicity and targeting special sites on the PI3K/Akt signaling. As the special marine environment provides the diversity of marine natural products to offer unique anticancer agents with promising clinical value, it is worthy to identify more anticancer agents that can be used clinically.

Acknowledgements

The study was supported by the National Natural Science Foundation of China (Nos. 81573457 and 81773776). We are also grateful to the support from the Taishan Talents Project of Shandong Province and the Department of Science and Technology in Shandong Province of China (Nos. ZR2017MH117, 2018YYSP025, and ZR2017MH 027), and Department of Science and Technology of Si- chuan Province, China (Nos. 2017HH0104 and 2019YFS 0116).

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June 16, 2020;

July 3, 2020;

December 25, 2020

© Ocean University of China, Science Press and Springer-Verlag GmbH Germany 2021

# The two authors contributed equally to this work.

. E-mail: xiukunlin@126.com

(Edited by Qiu Yantao)