Chemical Diversity of Soft Coral Steroids and Their Pharmacological Activities

The review is devoted to the chemical diversity of steroids produced by soft corals and their determined and potential activities. There are about 200 steroids that belong to different types of steroids such as secosteroids, spirosteroids, epoxy- and peroxy-steroids, steroid glycosides, halogenated steroids, polyoxygenated steroids and steroids containing sulfur or nitrogen heteroatoms. Of greatest interest is the pharmacological activity of these steroids. More than 40 steroids exhibit antitumor and related activity with a confidence level of over 90 percent. A group of 32 steroids shows anti-hypercholesterolemic activity with over 90 percent confidence. Ten steroids exhibit anti-inflammatory activity and 20 steroids can be classified as respiratory analeptic drugs. Several steroids exhibit rather rare and very specific activities. Steroids exhibit anti-osteoporotic properties and can be used to treat osteoporosis, as well as have strong anti-eczemic and anti-psoriatic properties and antispasmodic properties. Thus, this review is probably the first and exclusive to present the known as well as the potential pharmacological activities of 200 marine steroids.

In this review, we will look at rare and unusual steroids isolated from soft corals belonging to the order of Alcyonacea. The biological activity of many steroids has not been determined and we present the pharmacological activities detected experimentally and predicted based on the structure-activity relationships using the PASS (Prediction of Activity Spectra for Substances) software. PASS estimates the probabilities of several thousand biological activities with an average accuracy of about 96%. Probability of belonging to the class of "actives" Pa is calculated for each activity, providing the assessment of the hidden pharmacological potential of the investigated soft coral steroids [42].
96%. Probability of belonging to the class of "actives" Pa is calculated for each activity, providing the assessment of the hidden pharmacological potential of the investigated soft coral steroids [42].

Steroids Derived from the Genus Sinularia
Sinularia is a specific group of soft octocorals, a genus belonging to the family Alcyoniidae (class Anthozoa, phylum Cnidaria) and it includes about 175 actual species, 47 of which have been described only in the last quarter of a century [46,47]. Sinularia species are the most abundant corals in the entire Indo-Pacific region, especially in shallow water and dominate reef substrates [47].
Nowadays, many species of coral of the genus Sinularia are well adapted and cultured for biological and medical research and they are also an excellent source of many biologically active metabolites, including diterpenoids, unusual steroids and triterpenoids [15,19,38,40,48,49].
Secosteroid (127) with epoxide at C-5 and C-6 group from the Formosan soft coral Cespitularia hypotentaculata exhibited cytotoxicity against HT-29 cells [126]. The structures of the steroids  are shown in Figure 6 and the potential biological activities are shown in Table 5. Two steroids, 11-acetoxy-9,11-secosterols, pinnisterols E (128) and I (129) with a 1,4-quinone moiety, were discovered from the gorgonian coral Pinnigorgia sp. Both identified compounds reduced elastase enzyme release [127]. (22R,23S,24S)-Polyoxygenated steroid named hippuristerone A (130) has been isolated from a Taiwanese gorgonian I. hippuris [128,129]. A rare steroid derivative named griffinipregnone (131) has been isolated from the octocoral Dendronephthya griffin and showed anti-inflammatory activity [130]. An unusual hemiketal steroid, named cladiellin A (132) was isolated from the soft coral Cladiella sp. [131] and a similar steroid 23-keto-cladiellin A (133) was obtained from the monohydroxylated sterol fraction of soft coral Chromonephthea braziliensis [132]. The structures of the steroids  are shown in Figure 7 and the potential biological activities are shown in Table 8. An unusual pentacyclic hemiacetal sterol nephthoacetal (134) and acetyl derivative (135) were isolated from soft coral Nephthea sp. Compound (134) exhibited a significant inhibitory effect with EC 50 value of 2.5 µg/mL, while having low toxicity with LC 50 > 25.0 µg/mL. The in vitro cytotoxic activity of two compounds exhibited moderate cytotoxicity with IC 50 values of 12 and 10 µg/mL, respectively [133].
from the monohydroxylated sterol fraction of soft coral Chromonephthea braziliensis [132]. The structures of the steroids  are shown in Figure 7 and the potential biological activities are shown in Table 8. An unusual pentacyclic hemiacetal sterol nephthoacetal (134) and acetyl derivative (135) were isolated from soft coral Nephthea sp. Compound (134) exhibited a significant inhibitory effect with EC50 value of 2.5 µg/mL, while having low toxicity with LC50 > 25.0 µg/mL. The in vitro cytotoxic activity of two compounds exhibited moderate cytotoxicity with IC50 values of 12 and 10 µg/mL, respectively [133].     Extract of the soft coral Dendronephthya gigantea demonstrated the antiproliferative effect against the proliferation of HL-60 human leukemia cells and MCF-7 human breast cancer cells. The steroid 12-hydroxy-16,17-dimethyl-pregn-4-ene-1,20-dione (136) was isolated from the coral sterol fraction [134].
Krempenes A (137) and B (138) are unprecedented pregnane-type steroids that have been isolated from the marine soft coral Cladiella krempfi [135]. Steroid (137) contains a very unusual structural motif, with a hexacyclic oxadithiino unit fused to the steroidal ring A.
Unusual steroid thioesters, parathiosteroids A (155) and C (156) were isolated from the 2-propanol extract of the soft coral Paragorgia sp. collected in Madagascar. Both compounds displayed cytotoxicity against a panel of three human tumor cell lines at the micromolar level [147].
The soft coral Lobophytum michaelae that lives on the coast of Taitung and its ethyl acetate extract contained three cytotoxic polyoxygenated steroids called michosterols A-C (157-159) [148] and also ethyl acetate extract the gorgonian Leptogorgia sp. collected from the South China Sea contained dihydroxy-ketosteroid (160) [149]. The structures of the steroids (157-177) are shown in Figure 8 and the potential biological activities are shown in Table 9.
Mar. Drugs 2020, 18, x 26 of 39 Figure 9. Bioactive steroids derived from soft corals. The I. hippuris, as well as other species of this genus, belong to the bamboo corals that live in the Central-West Pacific, the Indian Ocean, as well as in the Red Sea [161]. Studies of these corals in the last quarter of a century have shown that they satisfy their metabolic requirements for carbon through the products of photosynthesis [162] and only about 9% from bacterioplankton. These corals synthesize many metabolites, which are highly oxidized products that are oxidized by their own endogenous oxygen. These include numerous highly oxygenated and spiroketal steroids, many of which demonstrate anticancer activity against many cancer cells [163][164][165][166][167][168][169][170].

Comparison of Biological Activities of Natural Soft Coral Steroids
The biological activity of the molecule depends on its structure, which allows analyzing the structure-activity relationships (SAR). This idea was first proposed by Brown and Fraser in 1868 [172] more than 150 years ago; however, it was further developed in the mid-1970s [173,174].
The quantitative structure-activity relationships (QSAR) paradigm was first implemented in toxicology, pharmaceutical and medicinal chemistry, and, ultimately, various aspects of organic and bioorganic chemistry [175]. For over 50 years, the QSAR paradigm has been widely used due to its original postulate that activity was a function of the structure described by electronic attributes, hydrophobicity or steric properties [176]. The rapid and extensive development of methodologies and computational methods led to the definition and refinement of many approaches that introduce the paradigm into the practice of research and development [177].
Several computer programs can estimate with some degree of certainty the pharmacological activities of organic metabolites isolated from natural sources or synthetic compounds [178][179][180]. Classical (Q)SAR methods are based on the analysis of (quantitative) structure-activity relationships for a single or several biological activities using the compounds belonging to the same chemical series as the training set [181].
Computer program PASS, which is continuously updating and improving for the past thirty years [182], is based on the analysis of a heterogeneous training set included information about more than a million known biologically active compounds with data on ca. 10,000 biological activities [183,184]. Chemical descriptors implemented in PASS, which reflect the peculiarities of ligand-target interactions and original realization of the Bayesian approach for elucidation of structure-activity relationships provide the average accuracy and predictivity for several thousand biological activities equal to about 96% [185,186]. In several comparative studies, it was shown that PASS outperforms in predictivity some other recently developed methods for estimation of biological activity profiles [187,188]. Freely available via Internet PASS Online web-service [189] is used by more than twenty thousand researchers from almost a hundred countries to determine the most promising biological activities for both natural and synthetic compounds [184,186,[190][191][192][193]. To reveal the hidden pharmacological potential of the natural substances, we are successfully using PASS for the past fifteen years [194][195][196][197][198].
In the current study, we obtained PASS predictions for two hundred steroids produced by soft corals. PASS estimates are presented as Pa values, which correspond to the probability of belonging to a particular class of "actives" for each predicted biological activity. The higher the Pa value is, the higher is the confidence that the experiment will confirm the predicted biological activity [186].
We have selected about 200 steroids, of which 50 belong to the genus Sinularia, which represent different types of steroids such as secosteroids, spirosteroids, epoxy-and peroxy-steroids, steroid glycosides, halogenated steroids and steroids containing sulfur or nitrogen heteroatoms. The types of steroids presented represent the chemical diversity of these secondary metabolites. Therefore, the pharmacological activities of various types of steroids are of great interest.
Analyzing the data obtained using PASS, we can state that almost all steroids presented in this article exhibit potential anti-tumor activity with varying degrees of reliability. In addition, forty-one steroids demonstrate anti-tumor and related activity with a confidence level of more than 90 percent, which is of significant interest to the pharmaceutical industry. Figure 10 shows the distribution of steroids with antitumor and related activities in the corals of the genus Sinularia. This group of steroids has a high degree of certainty over 90 percent. We highlighted other activities in a separate column and named them Lipid metabolism regulators, which include such properties of steroids as anti-hypercholesterolemic, treatment of atherosclerosis, cholesterol synthesis inhibitor and hypolipemic activity. This group includes 32 steroids that show anti-hypercholesterolemic activity with over 90 percent confidence. It is known that hypercholesterolemia and oncogenesis are interrelated, as shown in many studies [199][200][201][202][203]. Therefore, these data are also of great practical interest in identifying the etiology of cancer and its treatment. The column additional activity presents the activities that steroids demonstrate and some activities can also be attributed to the main ones since the reliability of the activities of some steroids exceeds 90 percent of the reliability. For example, the crassarosteroside A (5) water-soluble steroidal glycoside isolated from the Soft Coral S. crassa, according to the authors of the article, demonstrated cytotoxicity against human liver carcinoma (HepG2 and HepG3) [51,52]. Our studies have shown that it demonstrates strong antitumor activity and can be successfully used for the treatment of proliferative diseases, in addition, it demonstrates anti-hypercholesterolemic activity and is also a respiratory analeptic. Figure 11 shows the predicted and calculated (log activities) pharmacological activities of crassarosteroside A. We highlighted other activities in a separate column and named them Lipid metabolism regulators, which include such properties of steroids as anti-hypercholesterolemic, treatment of atherosclerosis, cholesterol synthesis inhibitor and hypolipemic activity. This group includes 32 steroids that show anti-hypercholesterolemic activity with over 90 percent confidence. It is known that hypercholesterolemia and oncogenesis are interrelated, as shown in many studies [199][200][201][202][203]. Therefore, these data are also of great practical interest in identifying the etiology of cancer and its treatment. The column additional activity presents the activities that steroids demonstrate and some activities can also be attributed to the main ones since the reliability of the activities of some steroids exceeds 90 percent of the reliability. For example, the crassarosteroside A (5) water-soluble steroidal glycoside isolated from the Soft Coral S. crassa, according to the authors of the article, demonstrated cytotoxicity against human liver carcinoma (HepG2 and HepG3) [51,52]. Our studies have shown that it demonstrates strong antitumor activity and can be successfully used for the treatment of proliferative diseases, in addition, it demonstrates anti-hypercholesterolemic activity and is also a respiratory analeptic. Figure 11 shows the predicted and calculated (log activities) pharmacological activities of crassarosteroside A.
Steroids have also been found that exhibit rather rare and extremely specific activities. For example, steroids 18, 37 and 38 can be used to treat autoimmune diseases. Steroids 53, 54 and 161 show anti-osteoporotic properties and can be used to treat osteoporosis. Steroids 94 and 102 are immunomodulators that can be used in the treatment of patients with AIDS. Steroid 163 exhibited strong anti-eczematic and anti-psoriatic properties and steroid 125 exhibited anti-eczematic and spasmolytic properties. Steroid 142 is a hepato-protector, steroid 86 is an inhibitor of angiogenesis and steroid 87 can be used as a general anesthetic.

Conclusions
About 200 soft coral steroids are classified as different types of steroids such as secosteroids, spirosteroids, epoxy-and peroxy-steroids, steroid glycosides, halogenated steroids, polyoxygenated steroids and steroids containing sulfur or nitrogen heteroatoms. There also have been found steroids that exhibit rather rare and extremely specific activities. The PASS program is constantly evolving by increasing the database of both natural and synthetic compounds and increasing biological activities by incorporating experimental data. Currently, PASS contains over 1,000,000 chemical structures of natural and synthetic compounds associated with over 10,000 biological activities. These activities are included in the program and are taken from published articles, reviews and other official medical profile documents. Thus, analyzing the presented steroids, it can be stated that more than 40 steroids demonstrate antitumor and related activity with a confidence level of more than 90 percent. Another group, which consists of 32 steroids, demonstrates anti-hypercholesterolemic activity with more than 90 percent confidence.
In addition, ten steroids exhibit anti-inflammatory activity and 20 steroids can be classified as respiratory analeptic drugs. Some steroids with rare structures exhibit anti-osteoporotic properties and can be used to treat osteoporosis, as well as exhibit strong anti-eczemic and anti-psoriatic properties, exhibit anti-eczema and antispasmodic properties. Thus, these data show that soft coral  Table 1).
Steroids have also been found that exhibit rather rare and extremely specific activities. For example, steroids 18, 37 and 38 can be used to treat autoimmune diseases. Steroids 53, 54 and 161 show anti-osteoporotic properties and can be used to treat osteoporosis. Steroids 94 and 102 are immunomodulators that can be used in the treatment of patients with AIDS. Steroid 163 exhibited strong anti-eczematic and anti-psoriatic properties and steroid 125 exhibited anti-eczematic and spasmolytic properties. Steroid 142 is a hepato-protector, steroid 86 is an inhibitor of angiogenesis and steroid 87 can be used as a general anesthetic.

Conclusions
About 200 soft coral steroids are classified as different types of steroids such as secosteroids, spirosteroids, epoxy-and peroxy-steroids, steroid glycosides, halogenated steroids, polyoxygenated steroids and steroids containing sulfur or nitrogen heteroatoms. There also have been found steroids that exhibit rather rare and extremely specific activities. The PASS program is constantly evolving by increasing the database of both natural and synthetic compounds and increasing biological activities by incorporating experimental data. Currently, PASS contains over 1,000,000 chemical structures of natural and synthetic compounds associated with over 10,000 biological activities. These activities are included in the program and are taken from published articles, reviews and other official medical profile documents. Thus, analyzing the presented steroids, it can be stated that more than 40 steroids demonstrate antitumor and related activity with a confidence level of more than 90 percent. Another group, which consists of 32 steroids, demonstrates anti-hypercholesterolemic activity with more than 90 percent confidence.
In addition, ten steroids exhibit anti-inflammatory activity and 20 steroids can be classified as respiratory analeptic drugs. Some steroids with rare structures exhibit anti-osteoporotic properties and can be used to treat osteoporosis, as well as exhibit strong anti-eczemic and anti-psoriatic properties, exhibit anti-eczema and antispasmodic properties. Thus, these data show that soft coral steroids are very interesting in terms of their medical use. However, this requires additional extensive research.