Oceans as a Source of Immunotherapy

Marine flora is taxonomically diverse, biologically active, and chemically unique. It is an excellent resource, which offers great opportunities for the discovery of new biopharmaceuticals such as immunomodulators and drugs targeting cancerous, inflammatory, microbial, and fungal diseases. The ability of some marine molecules to mediate specific inhibitory activities has been demonstrated in a range of cellular processes, including apoptosis, angiogenesis, and cell migration and adhesion. Immunomodulators have been shown to have significant therapeutic effects on immune-mediated diseases, but the search for safe and effective immunotherapies for other diseases such as sinusitis, atopic dermatitis, rheumatoid arthritis, asthma and allergies is ongoing. This review focuses on the marine-originated bioactive molecules with immunomodulatory potential, with a particular focus on the molecular mechanisms of specific agents with respect to their targets. It also addresses the commercial utilization of these compounds for possible drug improvement using metabolic engineering and genomics.


Introduction
Immune system dysfunction leads to the development of allergies, autoimmune and chronic inflammatory diseases, and cancers. Inflammation has been suggested to be the principal cause of chronic illnesses such as obesity, diabetes, cancer, rheumatoid arthritis (RA), neurodegenerative, and autoimmune diseases (ADs). The hallmark of autoimmunity is chronic inflammation that leads to the release of pro-inflammatory cytokines and other mediators, known as danger-associated molecular patterns (DAMPS), that activate pathogen recognition receptors (PRR) expressed by immune cells. Autoantibodies recognize these DAMPs and activate myeloid immune cells with an enhanced inflammatory response, leading to exacerbation of the condition. This self-perpetuating cycle continues, in order to assist with injury repair [1][2][3].
Recent estimates suggest that 7.6-9.4% of the world's population is affected by immune-mediated diseases. Such diseases include inflammatory bowel disease (IBD), type 1 diabetes mellitus (TIDM), and RA. Women are up to ten times as likely to be sufferers as men [4]. ADs are among the ten leading causes of death for women, and affect them disproportionately in every age group up to 75 years of age [5]. According to the NIH report, around 23.5 million Americans have ADs, while the American Autoimmune Related Diseases Association (AARDA) puts this figure at 50 million. More than 80 different ADs have been identified and 40 additional diseases are suspected as being ADs [6,7]. The holy grail of immunotherapy is the reprograming of the immune system to maintain or restore homeostasis, and there is an urgent need to develop such drugs.
The search for novel immunomodulators is challenging, despite the existence of considerable amounts of demographic and epidemiological data about ADs. Questions about how autoimmunity is triggered and how self-tolerance is broken down remain to be fully answered. Nevertheless, progression polyphosphate mediated septic responses and hyper-permeability through the inhibition of p38 mitogen-activated protein kinases (MAPKs) activation. Downregulation of tumor necrosis factor (TNF-α), IL-6, NF-κB and ERK1/2 was also observed after administration of these three compounds [22]. A novel exopolysaccharide (EPS) EPS1-T14, a water-soluble non-toxic exopolymer obtained from the marine bacterium Bacillus licheniformis, is able to stimulate an immune response. EPS1-T14 exhibits antiviral activity, as it inhibits the replication of HSV-2 in human peripheral blood mononuclear cells (hPBMCs). EPS1-T14 also stimulates the Th1 cell-mediated immune response [23]. Another EPS, TA-1, isolated from the thermophilic marine bacterium, Thermus aquaticus, is the strongest candidate for the EPS-binding receptor such as toll-like receptors (TLRs). TA-1 stimulates the release of the proinflammatory cytokines TNF-α and IL-6 from murine macrophages via a TLR-2 mediated pathway [24]. Prodigiosin (Figure 1, 1) derived from marine bacteria such as Pseudoalteromonas denitrificans, Vibro psychroerythrus, and Serratia marcescens, has a strong inhibitory effect on many protozoan, fungal, and bacterial species, and induces apoptosis in cancer cell lines, as observed by the development of characteristic DNA laddering and apoptotic bodies [25,26]. Cycloprodigisin, a stable analog of prodigiosin (Figure 1, 1) from Pseudoalteromonas dentrificans inhibits TNF-α induced NF-κB activation, as determined through luciferase assay. This NF-κB-inhibitory effect of cycloprodigiosin was retained under multiple stimuli in HeLa, U373, and COS7 cell lines [27][28][29]. Some representative immunomodulatory and anti-inflammatory chemical constituents isolated from marine bacteria are listed in Table 1.

Sponges
Sponges are currently the most important source of biologically active natural marine products and are considered to be a treasure trove of drugs [53,54]. Due to their lack of physical defenses, they have evolved a wide suite of defensive chemicals to deter predators [55]. New biomolecules discovered from marine sponges have strong immunosuppressive activities (Table 3). Didemnins, members of a depsipeptide class of compounds isolated from the Caribbean tunicate Trididemnum solidum, exhibit a variety of biological activities [56]. In particular, didemnin B (Figure 1, 4) is characterized as immunosuppressive, and inhibits lymphocyte activation [57,58]. Target deconvolution studies, which aim to identify the molecular targets of active hits, have revealed that didemnin B (Figure 1, 4) binds to the eukaryotic elongation factor 1α and palmitoyl-protein thioesterase 1. Large amounts of didemnin B (Figure 1, 4) were taken up by proliferating cells, so this compound appears to be a promising drug for cancer treatment or the suppression of activation of the immune system [59].
In several other non-lymphoid cell lines (+)-discodermolide exhibited antiproliferative effects by arresting the cell cycle at G2 and M phase due to microtubule network stabilization [107,108]. Petrosaspongiolide M (Figure 1, 7) isolated from the Caledonian marine sponge Petrosa spongia nigra significantly inhibits chronic inflammation in rats and mice by diminishing eicosanoids and TNF-α production [88]. Petrosaspongiolide M (Figure 1, 7) decreases the NF-κB-DNA binding in response to zymosan in mouse peritoneal macrophages [109]. A marine sesterterpene, heteronemin ( Figure 1, 8), isolated from Hyritos sponge species has been found to affect cellular processes including cell cycle and apoptosis, and inhibits TNF-α-induced NF-κB activation [81]. Methanolic extracts of Xestospongia testudinaria, the Red Sea marine sponge, prevent carrageenan-induced acute local inflammation in rats. Malondialdehyde and NO in inflamed rat paws was decreased by this extract, while glutathione, glutathione peroxidase, and catalase activities were increased. It appears to have antioxidant, anti-inflammatory, and immunomodulatory effects [82]. Some representative immunomodulatory and anti-inflammatory chemical constituents isolated from marine sponges are listed in Table 3.

Marine Fungi
Recent developments in marine mycology have led to a large amount of research into natural products from substrate-insulated fungi in various marine habitats [121]. The discovery rate of novel marine-derived natural products from fungi increased exponentially over the period from 1970 to 2010 [121]. Cyclosporins (Figure 1, 12) are produced by species of fungi including Tolypocladium inflatum gams [122], Neocosmospora vasinfecta, Verticillium spp. and Microdochium nivale [123,124]. Owing to their potent immune-modulation properties, cyclosporins ( Figure 1, 12) are used in patients with organ transplants. The agent specifically binds to cyclophilin expressed in T lymphocytes. The production of IL-2, IL-3, IL-4, granulocyte colony-stimulating factor (G-CSF) and TNF-α is reduced by cyclosporine in T-lymphocytes [125,126]. Sirolimus (Figure 1, 13) (Rapamycin) a macrocyclic lactone immunosuppressive drug was also derived from the fungus Streptomyces hygroscopicus [127]. It binds to FK-bound protein 12 and serine threonine kinase, mTOR, inhibiting the transduction of IL-2R and other cytokine signals relevant to allograft rejection [128,129]. Endarachne binghamiae Polysaccharides (Sodium alginate, alginic) Stimulates concentration-dependent proliferation of T cells and significant induction of the production of TNF-α and nitric oxide in macrophages and IFN-γ in T cells. [130] Caulerpa cupressoides, Pterocladiella capillacea and Solieria demonstrate Lectins Improves the IL-10 induction and induces the immune response of Th2 in mouse splenocytes. [120] Gracilaria verrucosa Enone fatty acids Inhibits the production of NO, TNF-α, and IL-6 inflammatory biomarkers. [119] Sargassum ilicifolium Terpenes, steroids and lipids Demonstrate chemotactic, phagocytic and intracellular killing of human neutrophils, and show a significant immunostimulatory effect in vivo. [90,131] Laminaria japonica Laminarin oligosaccharides & polysaccharides Apoptotic cell death protein was significantly reduced by laminarin oligosaccharides. [132] Nannochloropsis oceanica Ethanol extract Inhibits NO generation and downregulates NF-κB and β-secretase activities in BV-2 cells. [133] Monostroma nitidum Sulfated polysaccharides RAW 264.7 cells were stimulated by polysaccharides, which produced considerable NO, and PGE2 induces strong immunomodulation. [134] Hijikia fusiforme Polysaccharides Enhanced activity for the proliferative response of spleen cells in endotoxin nonrespondent C3H / HeJ mice. [135] Gyrodinium impudicum Polysaccharides Gyrodinium impudicum show immunostimulatory effects and enhance the tumoricidal activities of macrophages and NK cells in vivo. [136] Ulva fasciata Lipopolysaccharides Ulva in the diet significantly increases defense factors such as haemogram, agglutination index, phagocytic rate, bacterial clearance and serum bactericidal activity. [92] Sargassum thunbergii Fucoidan Fucoidan enhances phagocytosis and macrophage chemiluminescence. [137] Meristotheca papulosa Polysaccharides Extracts of M. papulosa significantly stimulated the proliferation of human lymphocytes. [138] Focellatus Carrageenan λ-carrageenan showed antitumor activity and lymphocyte activation in mice transplanted tumor. [139] Chlorella stigmatophora Polysaccharides Chlorella stigmatophora extract shows anti-inflammatory effect in paw oedema test and immunomodulatory effects in delayed hypersensitivity test. [140]
Apart from their economic importance, fungi have been utilized as food, usually collected from their fruiting bodies, mushrooms. Some mushrooms can stimulate the immune system, modulate cellular and humoral immunity, and potentiate anti-tumorigenic activity, and potentially rejuvenate immune systems weakened by the chemotherapy and radiotherapy used for cancer treatment. This ability of mushrooms therefore qualifies them as candidates for immunotherapy in cancer and other diseases [168].

Mangroves and Other Higher Plants
Partially submerged in the ocean, mangroves form a tangled network of above-ground roots, which creates a unique and complex habitat for all sorts of marine life. Mangroves have long been used in fisher-folk medicine to treat disease [169,170]. Some mangroves, like Rhizophora mangle and R. mucronata, have been screened for their anti-ulcer, anti-viral, and anti-inflammatory activities [171][172][173]. Leaf extract from Rhizophora apiculata has been shown to inhibit HIV-1 or HIV-2 or SIV viruses in various cell cultures [174]. Extract of Rhizophora apiculata has shown anti-inflammatory and anti-tumor activity against B16F10 melanoma cells in BALB/c mice. Rhizophora apiculata substantially reduces acute inflammation in mice induced by carrageenan, as well as inflammatory oedema induced by formalin [175]. Extract of rhizome from Acorus calamus inhibited the growth of many human and mouse cell lines. In hPBMCs the production of IL-2, NO, and TNF-α was inhibited, IFN-γ and cell-surface markers CD16 and HLA-DR were not affected, but CD25 was downregulated [176]. Some representative immunomodulatory and anti-inflammatory chemical constituents isolated from marine mangroves are listed in Table 6.

Marine Animals and Others
Entire marine animals, and their parts, contribute to the triggering of several biomedical mechanisms involved in inflammatory/allergic cascades [177]. An extract from the Caribbean Gorgonian Pseudopterogorgia elisabethae shows anti-inflammatory activity due to the presence of unusual diterpene glycosides, and is now used in cosmetic skin products as an anti-allergic factor [178]. Stichodactyla toxin (ShK)-186, a peptide toxin from sea anemones, blocks Kv1.3 potassium channels with a high degree of specificity. Kv1.3 potassium channels play a critical role in regulating the function of effector-memory T cells and class-switched memory B cells that are implicated in ADs [179]. Whole-body extracts of the marine prawn Nematopaleamon tenuipes (PEP), two gastropods, Euchelus asper (EAE) and Hemifusus pulgilinus (HPE), produced immunosuppression on Swiss albino mice in a concentration dependent manner [180]. An α-d-Glucan called MP-A, isolated from Mytilus coruscus (hard-shelled mussel), has shown anti-inflammatory activity in THP-1 human macrophage cells. MP-A suppresses LPS-induced TNF-α, NO, and PEG2 production via the TLR4 pathway [181]. Fatty acid extract from the tunicate Halocynthia aurantium significantly and dose-dependently increases NO and PGE2 production in RAW264.7 cells, producing immune enhancement without cytotoxicity. These fatty acids also regulate the transcription of immune-associated genes, including iNOS, IL-1β, IL-6, COX-2, and TNF-α [182]. Some representative immunomodulatory and anti-inflammatory chemical constituents isolated from marine creatures are listed in Table 6. Chaetomium globosum Chaetoglobosin Fex Suppresses LPS-stimulated IL-6, monocyte chemotactic protein-1, and TNF-α in peritoneal macrophages and mouse macrophage cells. [188] Penicillium paxilli Ma(G)K Pyrenocine A Inhibits gene expression in LPS-stimulated macrophages due to NF-κB-mediated signal transduction. [189] Ecklonia stolonifera Phlorofucofuroeckol (Phlorotannin) Inhibits NO and PGE2 production by the suppressing iNOS and COX-2 protein expression. [190] CD, cluster of differentiation; COX, cyclooxygenase; IL, interleukin; JNK, c-Jun NH2-terminal kinase; iNOS, inducible nitric oxide synthase; LKB4; leukotriene B4; MAPK; mitogen-activated protein kinase; MHC, major histocompatibility complex; NF-kB, nuclear factor-κB; NO, nitric oxide; PGE2, prostaglandin E2; PLA2, phospholipase A2; TLR, toll-like receptor; TNF, tumor necrosis factor. Table 6. Mangroves, corals and other marine creatures and their therapeutic chemical constituents.

Anti-inflammatory and Immunomodulatory Effects of the Chemical Constituents of Marine Flora
Marine organisms are not only adapted to life in water with high salt concentrations, but have incorporated halogens into their chemical constituents, since ocean water contains chloride, bromide and iodide [194]. The extensive utilization of halogen ions by various marine organisms has important consequences for their overall composition. A plethora of chemical compounds have been discovered from this source. The chemical uniqueness of marine organism-derived compounds has accelerated drug discovery from those marine sources which have the highest probability of having novel molecules and interesting biological activity [236]. Marine flora is a prolific source of bioactive constituents including polysaccharides, oligosaccharides, terpenoids, steroids, alkaloids, polyphenols and antioxidants.

Polysaccharides
The most abundant and chemically complex organic molecules in the oceans are polysaccharides (Figure 1, 19) [237]. Polysaccharides (Figure 1, 19) of marine origin are a class of biochemical compounds that has been shown to have valuable therapeutic properties. These compounds are considered to be biocompatible and to have little or no toxicity. Marine algae and bacteria possess an extensive and valuable chemical library of unique polysaccharides. Sulphated polysaccharides can enhance the innate immune response by promoting the tumoricidal activities of macrophages and natural killer cells [136,[238][239][240]. Polysaccharides (Figure 1, 19) from macroand microalgae have anti-inflammatory and immunomodulatory properties [241,242]. Among the marine algal polysaccharides, fucoidans, which are fucose-containing sulphated polysaccharides from brown seaweeds, have immunomodulatory effects [243,244]. EPCP1-2, a marine EPS extracted from Crypthecodinium cohnii, exhibits anti-inflammatory activity achieved by regulating the TLR4 pathway [245]. P-KG03 sulphated polysaccharide, derived from marine microalgae Gyrodinium impudium strain KG03, activates the production of NO in a JNK-dependent manner and stimulates the production of cytokines IL-1β and 6 and TNF-α in macrophages, and prevents the growth of tumor cells in vitro and in vivo [136,150]. Alginic acid, a colloidal polysaccharide from brown seaweed, inhibits the secretion of TNF-α and IL-1 [246]. Several bacteria found in the deep-sea, in shallow hydrothermal vents, the Antarctic, and hypersaline lakes produce EPS [247][248][249][250][251]. EPS1 from a haloalkaliphilic, thermophilic strain of Bacillus licheniformis T14 hinders HSV-2 replication in hPBMCs. The non-cytotoxic exopolymer EPS1-T14 can stimulate the immune response and thus contribute to host defense against viruses [23].

Alkaloids
Alkaloids, a structurally diverse group of secondary metabolites containing nitrogen, have a range of biological activities. Alkaloids are mostly found in higher plants, but many marine organisms also contain alkaloids [252,253]. Alkaloids from the marine sponges Axinella verrucosa and Acanthella aurantiaca have been characterized as NF-κB-specific inhibitors [100]. An oxindole alkaloid, convolutamydine A (Figure 1, 20) and its two analogs, ISA003 and ISA147 from marine bryozoans inhibits the formalin-induced licking behavior significantly in mice, migration of leucocytes, and expression of COX-2, PGE2, iNOS, IL-6, and TNF-α in RAW 264.7 cells [196]. Neoechinulins A (Figure 1, 21) and B, two diketopiperazine indole alkaloids from marine fungus Eurotium sp. SF-5989 exert in vitro anti-inflammatory activity on LPS-stimulated RAW 264.7 cells. Neoechinulin A (Figure 1, 21) was considered safe in vitro using a cell viability assay, but neoechinulin B exhibited toxicity. Neoechinulin A (Figure 1, 21) derived from Microsporum sp. downregulates the formation or expression of COX-2, PGE2, NO, ROS, iNOS, IL-1, IL-6, and TNF-α in oligomeric amyloid-β activated BV-2 microglial cells. This compound also downregulates apoptosis mediated by activated microglia in pheochromocytoma PC-12 cells and reduces the nuclear translocation of NF-κB p50 and p56 subunits. Neoechinulin A (Figure 1, 21) can also inhibit neuroinflammation in Alzheimer's disease [254]. Cytochalasan-based alkaloid chaetoglobosin Fex (Cha Fex) (Figure 1, 22), isolated from the fungus Chaetomium globosum, suppresses IL-6, TNF-α and monocyte chemotactic protein-1 in LPS-stimulated peritoneal macrophages and RAW 264.7 cells. Cytokine mRNA expression is lowered, entry of the p65 subunit of NF-κB into the nucleus and LPS-elicited breakdown of IκBα is impaired, and the levels of the extracellular-signal-related kinase (ERK1/2), p38, and c-Jun is reduced by Cha Fex (Figure 1, 22) alkaloid. In addition, the upregulation of membrane-associated CD-14 expression induced by LPS on RAW 264.7 cells and human monocytes was suppressed [188]. Alkaloids have been used by humans for a variety of purposes for more than 4000 years. Alkaloids and alkaloid-containing taxa will undoubtedly continue to play an important role in modern drug development [255].

Polyphenols
More than 8000 polyphenolic compounds are found in marine flora, including phlorotannins, flavonoids, anthocyanins, tannins, lignin, epigallocatechin, epicatechin, catechin, and hydroxylated polybrominated diphenyl ethers [256][257][258]. Polyphenols with multiple phenolic structural units are bioactive and are widely distributed in plants [257] and have a wide range of biological activities including antioxidant [259], cardiovascular protective, anti-cancer [260,261], anti-inflammatory and immune-modulatory effects [262,263]. Modern molecular and cellular biology techniques have led to a greater understanding of the benefits arising from polyphenols [264][265][266][267]. Cellular signaling and regulation of gene expression by polyphenols through modulation of NF-κB has a significant impact on cancer and chronic inflammation. For instance, resveratrol acts on the NF-κB pathway at multiple levels and is able to down-regulate its expression, phosphorylation and transcription activity [268][269][270][271]. Diphlorethohydroxycarmalol (DPHC) (Figure 1, 23), a phlorotannin from Ishige okamuarae, exerts an anti-inflammatory effect by strongly inhibiting IL-6 production in LPS-stimulated RAW264.7 cells. In addition, DPHC inhibits the expression of signal transducer and activator of transcription 5 (STAT5) signaling and increases the production of suppressor of cytokine signaling 1 (SOCS1) [151]. A phlorotannin sub-fraction isolated from Fucus distichus reduces TNF-α, IL-10, MCP-1 and COX-2 expression. The sub-fraction also lowers downstream TLR activation and expression of inflammatory biomarkers. In view of the potential cellular signaling capabilities of polyphenols, phlorotannin is beneficial because it acts as a free radical scavenger, and can also modulate inflammatory signaling receptors such as TLRs and downstream protein pathways, including NF-κB, JNK and p38 MAPKs [152].

Steroids/Sterols
Steroids are lipophilic compounds derived from cholesterol, and have a variety of marine, terrestrial, and synthetic sources. Steroids and their metabolites play an important role in the physiology and biochemistry of living organisms. For example they are used as hormone antagonists [272], contraceptives [273], cardiovascular therapeutic agents [274], anti-cancer agents [275], osteoporosis treatments [276], anesthetics, antibiotics, anti-asthmatics, and anti-inflammatories [277].
Steroidal compounds isolated from sponges that modulate the pregnane X receptors (PXRs) are effective in reducing intestinal inflammation by manipulating NF-κB activity [278,279]. Solomonsterol A (Figure 1, 24) from the marine sponge Theonella swinhoei is a selective PXR agonist and has been shown to mitigate systemic inflammation and immune system disturbances in a RA mouse model. Solomonsterol A (Figure 1, 24) inhibits the development of arthritis caused by anti-collagen antibodies in transgenic mice harboring a humanized PXR2. Solomonsterol A (Figure 1, 24) reduces the expression of the inflammatory markers TNF-α, IFN-γ and IL-17 and chemokines MIP1-α and RANTES, which reduces the inflammatory response [280]. Pregnane-type steroids derived from the soft coral Seleronephthya gracillimum show in vitro anti-inflammatory activity by reducing the accumulation of the pro-inflammatory proteins iNOS and COX-2. Sclerosteroid K (Figure 1, 25) and M (Figure 1, 27) reduce iNOS accumulation, while LPS-stimulated COX-2 accumulation in RAW 264.7 macrophages is prevented by these sclerosteriods [197]. Ergosta-7, 22-dien-3-ol (Figure 1, 28), an unsaturated sterol from the spiny sea star Marthasterias glacialis has anti-inflammatory activity on RAW 264.7 cells in vitro. The inflammatory markers NF-κB, iNOS, IL-6, and COX-2 are downregulated. Sterol ergosta-7, 22-dien-3-ol is most effective, but a potentially synergistic effect was obtained when this compound was administered with other compounds [198]. Steroids isolated from the starfish Astropecten polyacanthus, downregulate the generation of IL-6, IL-12 p40, and TNF-α in LPS-stimulated bone marrow-derived dendritic cells [199]. Polyoxygenated steroids michosterols A-C extracted from the soft coral Lobophytum micchaelae exhibit potent anti-inflammatory activity in stimulated neutrophils by suppressing the superoxide anion generation and elastase release in N-formyl-methionyl-leucyl-phenylalanine/cytochaslasine B (fMLP/CB) [200].

Miscellaneous Compounds with Anti-oxidant Activities
Antioxidants are agents which modulate the levels of highly reactive oxygen species (ROS), which cause damage by binding to biomolecules such as DNA. Antioxidants act by neutralizing ROS produced during biochemical reactions. In diseases such as Alzheimer's, Parkinson's, atherosclerosis, stroke, cancer, diabetes, RA, and IBD the antioxidant potential of bio-modulators has been studied and the underlying prophylactic and therapeutic aspects investigated [281][282][283][284][285][286]. Antioxidants can function as immune modulators and are used in conjunction with mainstream therapy in some diseases. Antioxidants have been used in disease prevention, where they serve as free radical scavengers. Tocopherols in humans and mice suppress PGE2 synthesis and enhance cell-mediated immunity [287][288][289]. Selenium augments the phagocytic abilities of macrophages and prevents CD8+ T lymphocyte damage [290]. Astaxanthin and fucoxanthin, marine carotenoids, appear to be biologically more effective than terrestrial carotenoids [291][292][293][294][295]. Astaxanthin decreases the production of NO, iNOS activity, and the production of PGE2 and TNF-α in RAW 264.7 cells in a dose-dependent manner [296]. Fucoxanthin prevents inflammation by inhibiting inflammatory cytokines TNF-α and IL-1, and limits the expression of COX-2 and iNOS, as well as eliminating excess ROS [297].
Vitamin C is an electron donor and acts as potent water-soluble antioxidant helping to prevent protein, lipid and DNA oxidation. Supplementation with vitamin C reduces IgE and histamine levels by increasing IFN-γ and deceasing IL-4, an observation which indicates the suppression of immune response of type Th2-type cytokines [298]. Trace elements such as iron, copper, selenium and antioxidants are found in eight species of red (Hypnea spinella, Gracilaria textorii, Gracilaria vermicullophyla), green (Caulerpa sertularioides, Codium simulans, Codium amplivesiculatum Ulva lactuca) and brown (Dictyota flabellata) microalgae [299]. Due to environmental conditions such as temperature and high levels of irradiation, microalgae and seaweeds have high levels of ROS, which seaweeds deactivate with their high intracellular amounts of antioxidant compounds such as polyphenols, phycobilins, carotenoids and vitamins [300].

Metabolic Engineering and Genomic Approaches for Marine Compounds
Marine compounds that are used as drugs in microorganisms, plants and animals are synthesized in small amounts and are difficult to obtain in large quantities. This is where metabolic engineering comes into play. Using metabolic engineering, we can model organisms using existing metabolic reconstructions to discover gene knockouts which could improve the yield of products of interest. Genome-scale metabolic constraint-based flux models have been constructed for Streptomyces coelicolor A3 strains with the aim of improving and optimizing the production of antibiotics [301]. The deletion of the gonCP gene in marine actinobacterium Streptomyces coniferous resulted in improved antitumor activity of two derivatives, PM100117 and PM100118 [302]. By combining metabolic engineering and mutagenesis, it was possible to produce a level of astaxanthin in Xanthophyllomyces dendrorhous that was 89 times higher than that of the wild-type strain [303]. Advances in the understanding of microbial metabolic pathways, together with the use of new combinatorial techniques and random mutagenesis, can increase product yield and enhance efficacy [304]. Genome-scale metabolic models have been developed for several non-marine organisms and several are currently used in industrial settings. Using genomic proteomic transcriptomic and other omics data across various conditions from in-vivo experiments and literature of marine organisms the genome-scale metabolic model can be developed to produce overproducing strains of a target organism (Figure 2) [305]. In many cases, the chemical structures of biomolecules can be predicted to a certain extent, based on the analysis and biosynthetic logic of the enzymes encoded in a biosynthetic gene cluster, and their similarity to known counterparts [306]. Genomic analysis provides new insights into marine biodiversity and can reveal new drug sources. Retrieval of genomic information from marine microorganisms can also be used for the discovery of new drug molecules from microorganisms that are yet to be cultivated [307]. The CRISPR-Cas9 gene editing technology has been used in large number of microorganisms for genome modulation [308]. Researchers assembled a multifunctional metabolic engineering system based on CRISPR, which makes use of an RNA-guided nuclease. This system allows metabolic engineers to modify microorganisms via gene modifications and substitutions, and provide a practical means to reduce metabolic flux through redundant metabolic pathways and direct energy towards production of the target compound [309].
the genome-scale metabolic model can be developed to produce overproducing strains of a target organism (Figure 2) [305]. In many cases, the chemical structures of biomolecules can be predicted to a certain extent, based on the analysis and biosynthetic logic of the enzymes encoded in a biosynthetic gene cluster, and their similarity to known counterparts [306]. Genomic analysis provides new insights into marine biodiversity and can reveal new drug sources. Retrieval of genomic information from marine microorganisms can also be used for the discovery of new drug molecules from microorganisms that are yet to be cultivated [307]. The CRISPR-Cas9 gene editing technology has been used in large number of microorganisms for genome modulation [308]. Researchers assembled a multifunctional metabolic engineering system based on CRISPR, which makes use of an RNAguided nuclease. This system allows metabolic engineers to modify microorganisms via gene modifications and substitutions, and provide a practical means to reduce metabolic flux through redundant metabolic pathways and direct energy towards production of the target compound [309].

Conclusions
Chemical substances derived from marine organisms have proven to be a very effective in the prevention and treatment of disease. The recent development of new marine-derived treatments for cancer, inflammation and infectious diseases suggest that a focus on the development of marine

Conclusions
Chemical substances derived from marine organisms have proven to be a very effective in the prevention and treatment of disease. The recent development of new marine-derived treatments for cancer, inflammation and infectious diseases suggest that a focus on the development of marine medicines could be very valuable. The discovery of novel chemicals with therapeutic potential from marine sources requires the exploration of unique habitats such as deep-sea environments, as well as isolation and culturing of marine microorganisms. The traditional pharmaceutical fit of marine microorganisms as model natural product drug sources makes them more attractive. The production of bulk quantities of microbe-derived drugs is still challenging and can be addressed using metabolic engineering, in order to meet the growing need for a wide range of pharmaceuticals. Marine organisms both known and as yet undiscovered, may hold answers to some of our most pressing medical problems.

Conflicts of Interest:
The authors declare that there are no conflict of interest.