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Appendix 1

Drugs from the sea

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MANOALIDE Molecular Weight: 418.566 g/mol Molecular Formula: C25H38O5 Source: Indo-Pacific sponge Luffariella variabilis (PORIFERA) Activity: anti-inflamatory, analgesia Status: Withdrawn from Phase II clinical trials. Manoalide was isolated from the sponge Luffariella variabilis collected in the Indo-Pacific. It is a member of a chemical family known as the sesquiterpenes. Although this natural product was originally reported as an antibiotic, follow-up work revealed manoalide possesses promising analgesic and anti-inflammatory properties. The compound works by inhibiting Phospholipase A2 (PLA2), which plays an important role in the inflammation process. Inhibitors of PLA2 like manoalide hold potential as treatments against conditions such as rheumatoid arthritis. Allergan Pharmaceuticals licensed manoalide and took it into Phase II human clinical trials as an experimental treatment for psoriasis. The company discontinued clinical trials, however, when it appeared that topical treatment did not allow sufficient quantities of the active drug to pass through the patient's skin. Allergan continues to pursue a synthetic medicinal chemistry program around manoalide. The compound is currently available as a commercial probe for PLA2 inhibition, but not as a therapeutic drug. PSEUDOPTEROSINS Source: Caribbean sea whip Pseudopterogorgia elisabethae (CNIDARIA) Activity: anti-inflammatory and analgesic agent Status: in use as a commercial skin cream additive; in preclinical development for medical applications The pseudopterosins were isolated from a Caribbean soft coral species called a sea whip (Pseudopterogorgia elisabethae). They belong to a class of compounds known as tricyclic diterpene glycosides. Pseudopterosins have been shown to possess

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potent anti-inflammatory and analgesic (pain relief) properties. They appear to work by inhibiting the synthesis of eicosanoids, (locally functioning hormone-like substances) in specific white blood cells called polymorphonuclear leukocytes. The extreme selectivity with which the pseudopterosins target their activity is intriguing to researchers. They appear to be pharmacologically distinct from other non-steroidal anti-inflammatory drugs (NSAIDs) and their mechanism of action appears to be novel as well. The pseudopterosins have been licensed to a pharmaceutical firm for medical use as anti-inflammatory drugs. At least one of the pseudopterosins has been brought through preclinical tests and an Investigational New Drug (IND) application has been filed with the U.S. Food and Drug Administration. A pseudopterosin extract has found its way to the non-pharmaceutical marketplace as an additive to prevent skin irritation in a line of Estèe Lauder cosmetic skin care products. ZICONOTIDE (Prialt®, SNX 111) Molecular Weight: 2639.16 g/mol Molecular Formula: C10H172N36O32S7 Source: Derivative of a conotoxide of cone snails Conus geographicus, Conus magus (MOLLUSCA). Activity: Analgesic Status: FDA-Approved Dec. 2004 for the management of severe pain (see below) Ziconotide is a synthetic derivative of short (25 amino acid) peptide extracted from the venom of predatory cone snails (Conus geographicus, Conus magus). This drug, a member of a newly described chemical family called the conopeptides, is a generating a lot of interest as a potential pain management drug. Results from clinical trials to date have suggested ziconotide's effectiveness in treating pain may be from fifty to several thousand times better than that of morphine. It is non-addictive and thus may be suitable for long-term use if it also proves to be non-damaging (but see below). Cancer and AIDS patients and people suffering from chronic neuropathic pain are the initial target patients for the drug. Ziconotide is also reportedly being administered intravenously in some surgery patients. Ziconotide appears to suppress pain by targeting and blocking specific neuron presynaptic ion channels (called N-type calcium channels) to short-circuit neurotransmitter release in nerves that transmit pain signals. The precisely targeted mode of action effectively blocks pain while still allowing the rest of the nervous system to function properly. This represents an important advantage of this drug over that of currently available opiates with more systemically suppressive effects (e.g. sedation, respiratory depression, etc.).

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The fact that ziconotide and other conotoxins are short (usually 20-30 amino acids) means that synthetic derivatives are typically easy to produce. The potential risk of neurotoxicity is being evaluated but the drug has been conditionally approved for use as the significant benefits of this powerful drug outweigh the risk. A synthetic version of the drug, SNX-111, is manufactured by licensee Elan Corporation under the trade name Prialt®. The drug achieved its primary endpoint in Phase III clinical trials in January 2003, and was approved in late 2004 in the US and early 2005 in the EU for the management of chronic pain in patients for whom intrathecal therapy (e.g., by means of an implanted infusion pump) is warranted, and who are intolerant of or resistant to other treatment. A similar synthetic conopeptide called AM336 31 has also been recently reported. This compound has also very recently been put into clinical trials. A natural conotoxin called ACV1, isolated from Australian C. victoriae, has also recently been placed into early clinical trials as a treatment for neuropathic pain associated with diabetes. ANABASEINE Molecular Weight (Sarcodictyin A): 160.216 g/mol Molecular Formula (Sarcodictyin A): C10H12N2 Source: Various nemertine worms, esp. Paranemertes peregrina (NEMERTEA) Activity: Anti-Alzheimer agent Status: Phase I clinical trials. Pioneering marine pharmacological work carried out by Bacq in the 1930s examined the toxic activity of extracts prepared from predatory marine nemertines. The worms use the substance as a venom to paralyze prey and it may also play a role in deterring predators. The state of the science was not yet advanced enough to permit purification and complete chemical characterization of the suspected toxin Kem dubbed "amphiporine." Later, in the early 1970s, William Kem extracted and successfully purified a hoplonemertine toxin with similar activity from the nemertine Paranemertes peregrina collected at Friday Harbor WA. Chemical characterization revealed that hoplonemertine toxin was structurally identical to the compound anabaseine, first synthesized in the laboratory in the 1930s and structurally related to a compound isolated from tobacco. The same compound has more recently been found as a venom component in two different ant species (Aphaenogaster spp.). Anabaseine is a nicotinoid alkaloid. It is capable of stimulating vertebrate neuromuscular nicotinic receptors and increasing cholinergenic transmission. As such it has potential as a treatment of cognitive function loss. A synthetic analog, DMXBA (GTS-21) has exhibited memory enhancing effects in recipients. The compound is currently under

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license by the Japanese pharmaceutical company Taiho and is in Phase I trials for treating Alzheimer's disease. DEBROMOHYMENIALDISINE Molecular Weight: 245.238 g/mol Molecular Formula: C11H11N5O2 Source: The Palauan sponge Stylotella aurantium (PORIFERA). Activity: anti-Alzheimer agent; treatment against osteoarthritis. Status: Phase I clinical trials. Debromohymenialdisine (DBH) is an alkaloid originally isolated from the shallow-water Palauan marine sponge Stylotella aurantium. The molecule is intriguing not just for its drug-like properties, but also because it's simple molecular structure has yielded to easy total synthesis in the laboratory. Research suggest the compound acts as a highly selective inhibitor of a specific target cell DNA damage checkpoint enzyme during the G2 phase of the cell cycle. The compound is a promising potential Anti-Alzheimer agent. It has been licensed to Genzyme Tissue Repair for commercial development as a possible treatment for osteoarthritis. The drug is reportedly in Phase I clinical trials. NEOVASTAT® (AE-941) Source: Taxonomically identified shark species harvested under sustainable conditions (CHORDATA) Activity: Anti-tumor agent; anti-angiogenic agent Status: Preclinical Neovastat (AE-941) is a derivative of shark cartilage extract. Rather than being a specific monomolecular compound, AE-941 is a defined standardized liquid extract comprising the < 500 kDa (kilodaltons, a unit of mass) fraction from shark cartilage. The multiple mechanisms of action thus far reported for AE-941 are impressive. As a primary MOA, research suggests AE-941 inhibits the binding of Vascular Endothelial Growth Factor (VEGF) to its receptors. Normally, when VEGF is secreted by tumors it binds to target endothelial receptors and directs the profusion of new capillaries to supply the tumor with nourishment. By blocking the receptor sites, AE-941preempts the formation of the new blood supply the growing tumor needs to sustain itself. AE-941 also inhibits the metastatic cellular machinery the tumor normally uses to disrupt the extracellular matrix of the surrounding host tissue. Additionally, the drug appears capable of inducing endothelial cell specific apoptosis and also of inducing an increase in

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endothelial cell production of compounds that may cause the disintegration of blood vessels already present within the tumor. The anti-angiogenic and antitumor activity activity of AE-941 was first reported in 1997. It is now in Phase III trials in several countries for renal cell carcinoma and non-small-cell lung cancer. The literature also confirms the drug's efficacy in stabilizing tumor progression and relieving pain in metastatic prostate cancer patients. More recently, AE-941 has received attention as a possible treatment against metastatic breast cancer. The drug's anti-angiogenic bioactivity further suggests it could be a valuable agent for use in patients suffering from multiple myeloma and other hematologic (blood) diseases. One of the characteristics of multiple myeloma is bone marrow angiogenesis, and treatment with an anti-angiogenic agent like neovastat shows potential. From an ecological standpoint, commercialization and broadened chemotherapeutic use of AE-941 may be problematic. Although the drug extract is produced from only taxonomically identified shark species harvested under ostensibly sustainable conditions, long-term exploitation of already critically threatened natural shark populations should be avoided if at all possible. SQUALAMINE Molecular Weight: 627.963 g/mol. Molecular Formula: C34H65N3O5S. Source: The shark Squalus acanthus (CHORDATA) Activity: Anti-tumor agent; Anti-angiogenic agent. Status: Phase I/Phase II clinical trials; also sold as a non FDA-approved dietary supplement Squalamine is an aminosterol isolated from the stomach and liver of the spiny dogfish, Squalus acanthus, a common New England coastal shark species. When it was discovered in 1993, the compound was reported to exhibit broad-spectrum antibiotic activity. Squalamine was licensed to Magainin Pharmaceuticals (now Genaera Corporation) for development. It has progressed to Phase II clinical trials as part of a combination treatment for non-responding solid tumors, and as a primary treatment against ovarian cancer. Phase I prostate cancer trials were slated to commence some time in 2004 and there is data suggesting efficacy against the commonest and deadliest brain tumors as well. Of considerable interest is published evidence suggesting that squalamine exhibits anti-angiogenic activity under certain conditions (angiogenesis is the formation and differentiation of blood vessels). The speculation and hope of researchers is that squalamine starves tumors by preventing the typical proliferation of blood vessels they require for nourishment. If this is the case, a

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squalamine-derived anti-tumor drug may be particularly useful since its use should not select for drug resistence in treated cancers because the target of the drug is not the tumor itself but rather the blood vessels supplying the tumor. The exact method of action is being investigated, but squalamine appears similar to another shark-derived product, neovastat, in its ability to inhibit tumor production of Vascular Endothelial Growth Factor (VEGF) and other such growth factor signals. Additionally, squalamine appears capable of inducing endothelial cell inactivation and apoptosis through inhibition of integrin (specialized receptor protein) expression and the disruption of cytoskeletal formation. Because squalamine is not a protein it can be taken orally as a pill, unlike other anti-angiogenic drugs being evaluated which must be administered intravenously to ensure they are not destroyed by digestive enzymes. In addition to squalamine's potential as an anti-tumor drug, the compound's licensee Genaera Corporation has also reported very promising Phase I and Phase II clinical results when the drug is used for the treatment of vision problems relating to age-related macular degeneration. In these cases, he drug's anti-angiogenic activity appears to inhibit the choroidal neovascularization associated with this eye condition. CONTIGNASTEROL (IZP-94005, IPL576,092) Molecular Weight: 508.687 g/mol Molecular Formula: C29H48O7 Source: The sponge Petrosia contignata (PORIFERA) Activity: anti-asthma agent Status: In clinical trials (various phases) Although isolation of the natural product contignasterol (IZP-94005) from the sponge Petrosia contignata was first reported in the early 1990s, it took another ten years for the structural configuration of the compound to be elucidated. This molecule has proven to be a good starting point for semisynthetic chemists, and some of the novel compounds derived from contignasterol were shown to demonstrate activity as histamine blockers in in vitro rodent cell cultures. The natural compound itself was deemed a good anti-asthma drug candidate based on performance in in vivo animal models. One contignasterol derivative, named IPL576,092, shows promise as an oral asthma medication. It was been introduced into clinical trials by the drug company Aventis Pharma and completed Phase II studies in 2002. More recently, IPL576,092 has entered clinical trials as a treatment for diseases causing inflammation of the eyes and skin. Interest persists in other contignasterol analogs, and some of these may now be in clinical trials as well.

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CRYPTOPHYCINS Molecular Weight (Cryptophycin 1): 655.177 g/mol Molecular Formula (Cryptophycin 1): C35H43ClN2O8 Source: marine and non-marine cyanophytes; Dysidea arenaria (PORIFERA) Activity: tubulin interactive agent, antifungal, cytotoxin Status: Clinical trials of semi-synthetic cryptophycin 52 discontinued in 2002. This family of compounds was initially reported from a terrestrial source (the Nostoc sp. cyanophyte component of a Scottish lichen). Representative compounds have also been found in species of free-living marine and non-marine cyanophytes and, more recently, from the Japanese sponge Dysidea arenaria. The expressed bioactivity that was originally pursued was activity as an antifungal agent. Development of the compound for this purpose was not pursued beyond preliminary investigations when it was deemed too toxic for human use. More recently it has been reported that cryptophycin 1 is an inhibitor of tubulin assembly in cells, likely binding to tubulin at the so-called "peptide binding site" in a manner similar to that suspected for dolastatin 10 and the hemiasterlin derivative HTI-286. Lilly licensed the compounds from the University of Hawaii and Wayne State University, and utilized a semi-synthetic approach to build the modified natural product cryptophycin 52. This compound progressed through Phase I and toward Phase II human clinical trials before trials were discontinued in 2002. Cryptophycin 24 (arenastatin A) from the Okinawan sponge Dysidea arenari is reported to be a potent cytotoxin. LASONOLIDES Source: Gulf of Mexico deep-sea sponge Forcepia sp. (PORIFERA) Activity: anticancer; MOA under investigation Status: Preclinical The lasonolides are a series of marine natural products under investigation for the treatment of cancer. The compounds were isolated in 1994 by scientists from the Harbor Branch Division of Biomedical Marine Research from the sponge Forcepia sp. found in Gulf of Mexico deep-sea habitats. These compounds are very potent and show especially promising properties for the treatment of pancreatic cancer. They kill cancer cells in a different way than most other cancer drugs. The exact mode of action is not yet fully understood, and is an area of active research. In addition to anti-proliferative and antitumor properties, these novel macrolide compounds display antifungal activity as well.

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MANZAMINE A

Molecular Weight: 548.761 g/mol Molecular Formula: C36H44N4O Source: The sponge Haliclona sp. (PORIFERA) Activity: Demonstrated activity against malaria, tuberculosis, HIV, and others. Status: Preclinical

TOPSENTINS Molecular Weight (Topsentin B1): 342.351 g/mol Molecular Formula (Topsentin B1): C20H14N4O2 Source: The sponges Topsentia genitrix, Hexadella sp., and Spongosorites ruetzleri (PORIFERA) Activity: anti-inflammatory agent Status: Preclinical The Topsentins are a class of natural products that have been extracted from several sponge species subsequent to their initial isolation from

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the sponge Spongosorites ruetzleri. These compounds have been shown to have significant anti-inflammatory properties. Although the precise mode of action is not known, the compound has been reported capable of suppressing immunogenic as well as neurogenic (originating in nerve tissue) inflammation. The topsentins may hold promise as an arthritis medication or as additives in anti-inflammatory creams for the treatment of skin irritations. Early research suggests the compounds may also one day prove useful as treatments against colon cancer, Alzheimer's disease, and inflammatory bowel disease. Isolation of the topsentins from a number of unrelated sponge species strongly suggests that the target compounds may actually be produced by bacteria or other microorganisms living within the tissues of the sponges. Some success has been reported in preparing synthetic topsentin analogs that retain good bioactivity. Such efforts will hopefully obviate the need for large-scale collection of topsentin-containing deepwater sponges as a source of natural product if and when clinical trials are commenced. At the 9th International Symposium on Marine Natural Products (Townsville, July 1998), the researchers recently reported success in preparing a relatively simple synthetic bis-indole derivative that has good anti-inflammatory activity, thereby circumventing the difficult and expensive task of collecting the deep-water sponge. BENGAMIDES and Derivatives Bengamide A Molecular Weight: 598.812 g/mol Molecular Formula: C32H58N2O8 Bengamide B Molecular Weight: 584.785 g/mol Molecular Formula: C31H56N2O8 Source: Fijian sponges: unidentified Jaspidae, Jaspis cf. coriacea (PORIFERA) Activity: tumor growth inhibitor Status: Synthetic analog LAF389 withdrawn from Phase I clinical trials in 2002. Two novel seven-membered ring heterocycles, bengamide A and bengamide B, were reported in 1986 isolated from an as yet undescribed Fijian sponge belonging to family Jaspidae. Since this time, a number of additional compounds from the bengamide class have been isolated, most notably from the Fijian sponge Jaspis cf. coriacea. Bengamides A and B were initially reported to exhibit in vitro toxicity to larynx epithelial carcinoma cells, and to have antibiotic and anti-helminthic activity (against the nematode Nippostrongulus braziliensis) as well. A synthetic bemgamide analog, LAF389, was developed and shown to inhibit certain tumor growth, apparently by means of methionine aminopeptidase inhibition. The

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pharmaceutical company Novartis (then Ciba-Geigy) brought LAF389 into Phase I clinical trials before deciding to withdraw this compound in 2002. There is still ongoing research that is centered on the Bengamides and their derivatives. A report from late 2003 describes a proteomics-based investigation in which LAF389 was used to identify protein signaling pathways altered by bengamide addition. KRN7000 Molecular Weight: 842.323 g/mol Molecular Formula: C50H99NO8 Source: Analog derived from the sponge Agelas mauritianus. (PORIFERA) Activity: antitumor, immunostimulatory Status: Phase I clinical trials (Europe and Asia) A collaboration between employees of Japan's Kirin Brewery and the University of the Ryukyus published a report describing the first known isolation of a chemical class called the alpha-galactosylceramides from a natural source (the sponge Agelas mauritianus). These compounds, dubbed the agelasphins, were shown to exhibit antitumor and possible immunostimulatory activity. Further work revealed pronounced in vivo activity against B16 melanoma in mice. This work prompted the synthesis of a number of derived compounds, including KRN7000. Survival rates of mice inoculated with B16 and EL-4 cancer cells and then treated with the synthetic compound were increased significantly over those that did not receive the drug. Observation that KRN7000 markedly stimulates lymphocytic proliferation under certain conditions indicates the drug may be a powerful biological response modifier. Another animal study revealed KRN7000 significantly inhibited tumor growth and metastasis in the liver. KRN700 activity is unusual in that it appears to stimulate the production of natural killer T (NKT) cells in the body. These are blood plasma cells with cytotoxic anti-tumor activity that target lipids rather than proteins. Recent efforts, led by the Kirin researchers, have uncovered similar NKT cell activators in mammals. KRN7000 was placed into Phase I trials for cancer immunotherapy in Asia and Europe in 2001. SPISULOSINE (ES-285) Molecular Weight: 321 g/mol. Molecular Formula: C18H40ClNO. Source: The bivalve Spisula (=Mactromeris) polynyma (MOLLUSCA) Activity: Antitumor agent. Status: Currently in Phase I clinical trials

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Isolation of this natural product from the commercially harvested Arctic surf clam Spisula (=Mactromeris) polynyma by researchers from the Spanish PharmaMar group was first reported in 1999. The molecule shows promise as an antiproliferative (anti-tumor) agent, and has intriguing mechanism of action. Cultured cells treated with spisulosine exhibit a greatly altered morphology. Examination of the cytoskeleton of the cells reveals the change is due to a loss of actin stress fibers, (bundles of actin filaments believed appear and disappear in response to mechanical stimuli). PharmaMar is presently directing Phase I clinical spisulosine trials in Europe. The fact that the Arctic surf clam is a long-lived species (many 40+ year individuals exist in the wild populations) that is slow to reach reproductive maturity (at 5-8 years) will become an important factor if the decision is made in the future to expand commercial harvest to meet demand for supplies of spisulosine. Technologies emerging from current aquaculture efforts directed toward commercial production of the closely related Atlantic surf clam (S. solidissima) may be transferable to S. polynyma. so that a consistent, fast growing product can be developed for drug development purposes. ECTEINASCIDIN 743 (Yondelis®, ET-743) Molecular Weight: 761.84 g/mol. Molecular Formula: C39H43N30O11S. Source: The tunicate Ecteinascidia turbinata (CHORDATA). Activity: Anti-cancer agent via apoptosis induction. Status: Phase II Clinical Trials, European Orphan Drug Designation against soft tissue sarcoma (see below). Ecteinascidin 743 (ET-743; Etrabectedin) was isolated from the Caribbean sea squirt (Ascidia) Ecteinascidia turbinata, a common component of the intertidal/subtidal mangrove prop root community. It is classified as a tetrahydroisoquinoline alkaloid. Preclinical trials showed ET-743 was active against a range of tumor types in standard animal models. Subsequent human trials

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showed efficacy against advanced soft tissue sarcoma, osteosarcoma and metastatic breast cancers. In Phase I human clinical trials the drug showed effectiveness against advanced-stage breast, colon, ovarian and lung cancers, melanoma, mesothelioma and several types of sarcoma. Phase II studies with ET-743 are currently being conducted in the United States and four other countries. Research into the mode of action of ET-743 has revealed that binding of the drug to target cell DNA inhibits cell division and leads to apoptosis of cancer cells. Setting the drug apart from others that trigger 'cell suicide' mechanisms is the fact that ET-743 induces apoptosis only during active gene transcription. This makes actively dividing cancer cells more vulnerable to drug toxicity than normal cells because they exhibit greatly accelerated transcription and translation rates. Ecteinascidin 743 also may hold promise in keeping tumors from becoming resistant to chemotherapy. Ecteinascidin 743 interferes with the gene that produces P-glycoprotein, a membrane protein that confers drug resistance on cancer cells by actively transporting toxic compounds (like drug therapies) out of the cells. Such activity suggests that ET-743 may become important as a key ingredient in multi-drug 'cocktails' if it can prevent target cells from developing resistance to the other drug therapies. ET-743 has been co-developed under the trade name Yondelis® by the Spanish marine pharmaceutical company PharmaMar and Johnson & Johnson subsidiary Ortho Biotech. The drug is well along in Phase II clinical trials. The company reports that Yondelis is the first new drug in 20 years that is effective against soft tissue sarcoma (STS) a rare but pernicious form of cancer. In 2001, Yondelis obtained Orphan Drug Designation for the treatment of STS in Europe. APLIDINE (Aplidin®, Dehydrodidemnin B) Source: The tunicate Aplidium albicans (CHORDATA). Activity: Anti-cancer agent via apoptosis induction. Status: Phase II clinical trials. Aplidine (Dehydrodidemnin B), has was isolated from the Mediterranean tunicate Aplidium albicans. It was first reported in a 1991 patent application. In preclinical animal tests, Aplidine exhibited marked anticancer properties, outperforming the related compound didemnin B by a factor of six. Aplidine differs chemically from didemnin B and the other didemnins only in the structure of its side chain. This fact, along with the small size and relatively simple structure, has allowed research to achieve total synthesis of didemnin analogs. Synthesis has permitted the replacing of some amino acids non-essential for bioactivity with covalent bonds, which has increased resistance to enzymatic degradation and prolonged the

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bioactive lifetime. Analogs more active than any of the original natural didemnins have been produced in this manner. The molecule has been described as a multifactorial apoptosis inducer, and it has other beneficial attributes such as low toxicity and a high specificity for tumor cells. The varied inferred mechanisms of action thus far reported include rapid and persistent activation of apoptosis and interruption of the tumor cell cycle at the G1-G2. The compound also inhibits the expression of receptor proteins (ornithine descarboxylase) and the secretion of proteins (vascular endothelial growth factor) involved in growth and vascularization of certain tumor types. Synthetically derived Aplidine, under the PharmaMar trade name Aplidin®, is currently in Phase II trials for a variety of cancers, including melanoma, colorectal, renal, lung, head and neck (non-cerebral), pancreatic and medullary thyroid carcinomas. Additional Phase II trials against other forms of cancer are commencing. Aplidin was granted "orphan drug status" in the EU for acute lymphoblastic leukemia in 2003. HEMIASTERLINS Molecular Weight (Hemiasterlin): 526.711 g/mol Molecular Formula (Hemiasterlin): C30H46N4O4 Source: Sponges of genus Auletta, Siphonochalin (PORIFERA) Activity: cytotoxic and tubulin interactive agents Status: Preclinical This class of novel marine oligopeptides have been shown to be potent antitumor agents. Representatives of this class of natural products have been isolated from extracts prepared from sponges residing in two distinct genera (Auletta; Siphonochalina). Three different hemiasterlins with drug development potential (hemiasterlin, hemiasterlin A, hemiasterlin C) have been the subject of chemical and biological investigations. These molecules exhibit cytotoxic and antitubulin activity similar to that seen in the dolastatins. Mitotic inhibition occurs through binding to tubulin at the vinca/peptide region in a manner similar to dolastatin. The production of synthetic hemiasterlin analogs, facilitated by the simple tripeptide molecular structure, has been achieved. Studies have demonstrated that one analog, HTI-286, effectively inhibits tubulin polymerization, disrupts cell microtubule organization, and induces mitotic arrest and apoptosis. KAHALALIDE F Molecular Weight: 1477.87 g/mol Molecular Formula: C75H124N14O16

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Source: Hawaiian sacoglossan Elysia rufescens (MOLLUSCA) Activity: Cytotoxin; gene inhibitor Status: Currently in Phase II clinical trials Kahalalide F was isolated from a sacoglossan (sea slug) mollusc, Elysia rufescens, collected from Hawaii. Kahalalide F appears capable of disrupting lysosome [def] membranes within certain target cells, thereby initiating apoptosis (programmed cell death). The drug also appears to inhibit the expression of certain specific genes that are involved in DNA replication and cell proliferation, thereby inhibiting tumor spreading and growth. The drug shows promise in treating a broad range of tumors, including non-small cell lung cancer (NSCLC), melanoma, androgen-independent prostate cancer and hepatocellular carcinoma. Under development by the Spanish marine pharmaceutical company PharmaMar, Kahalalide F completed Phase I human clinical trials in patients, and entered into Phase II trials for non-small cell lung cancer (NSCLC) and in melanoma in July 2004. THIOCORALINE Molecular Weight: 1157.41 g/mol Molecular Formula: C48H56N10O12S6 Source: Micromonospora marina (Actinomycete bacteria) Activity: DNA polymerase ± inhibition Status: Preclinical Belonging to the chemical family known as the thiodepsipeptides, thiocoraline was first isolated from Micromonospora marina, an actinomycete bacterium collected from coastal Mozambique, southeast Africa. The Spanish marine drug company PharmaMar has reported that the compound shows activity against several standard drug screens, including breast cancer, colon cancer, renal cancer, and melanoma. Target cells appear to be inhibited through inhibition of DNA polymerase ± enzyme. The most recent published literature suggests thiocoraline is still undergoing advanced preclinical evaluation.

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BRYOSTATIN 1 Molecular Weight: 905.033 g/mol Molecular Formula: C47H68O17 Source: The bryozoan Bugula neritina (ECTOPROCTA) Activity: anti-cancer and immunosuppressive, based on protein kinase C binding inhibition Status: In Phase II clinical trials. This natural product was originally extracted from the bryozoan (a sessile, moss-like marine animal) Bugula neritina collected in the Gulf of California and Gulf of Mexico. More recent work has demonstrated that the compound is most likely produced by the microbial symbiont Endobugula sertula. This compound and other bryostatins produced by the microbial associate are exploited by the host as a chemical means of defense, particularly in the larval stage. Bryostatin 1 is a macrocyclic lactone that belongs to a diverse class of complex products called polyketides. The compound has demonstrated promising anti-cancer, anti-tumor, and immunostimulant activities that are apparently related to its ability to bind to protein kinase C, and enzyme involved in the up-regulating (switching on) and down-regulating (switching off) of certain proteins. In 2001, Bryostatin 1 was licensed from Arizona State University for commercial development by German pharmaceutical company GPC Biotech and is currently in several Phase II human clinical trials under the guidance of the National Cancer Institutes. That same year, GPC Biotech also struck a licensing deal with Stanford University to develop and commercialize synthetic analogs of Bryostatin 1, known as Bryologs. The most recent trials have shown that Bryostatin enhances the effectiveness of existing chemotherapies such as taxol and cisplatin, but is relatively ineffective on its own. If Bryostatin proceeds to the development phase, it will likely be developed as a tandem treatment for breast, ovarian and lung cancers and others that respond somewhat to existing regimens. It has an advantage over some current chemotherapies that inhibit red blood cell production and thus require blood transfusions. Bryostatin, in contrast, appears to stimulate red blood cell production. Bryostatin was granted orphan drug status for use in combination with Taxol for the treatment of esophageal cancer by the FDA in december of 2001. Similar status designation in the EU was awarded in 2002. Despite the apparent progress, in 2003 GPC Biotech made a strategic decision to discontinue its bryostatin development programs, stating that trials "had not provided sufficient evidence of efficacy combined with an acceptable toxicity profile to move the drug candidate forward."

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DEBROMOHYMENIALDISINE Molecular Weight: 245.238 g/mol Molecular Formula: C11H11N5O2 Source: The Palauan sponge Stylotella aurantium (PORIFERA). Activity: anti-Alzheimer agent; treatment against osteoarthritis. Status: Phase I clinical trials. Debromohymenialdisine (DBH) is an alkaloid originally isolated from the shallow-water. Palauan marine sponge Stylotella aurantium. The molecule is intriguing not just for its drug-like properties, but also because it's simple molecular structure has yielded to easy total synthesis in the laboratory. Research suggest the compound acts as a highly selective inhibitor of a specific target cell DNA damage checkpoint enzyme during the G2 phase of the cell cycle. The compound is a promising potential Anti-Alzheimer agent. It has been licensed to Genzyme Tissue Repair for commercial development as a possible treatment for osteoarthritis. The drug is reportedly in Phase I clinical trials. Girolline (Girodazole) Molecular Weight: 190.631 g/mol Molecular Formula: C6H11ClN4O Source: First reported from the sponge Pseudaxinyssa (= Cymbastela) cantharella (PORIFERA). Activity: protein synthesis inhibitor Status: Clinical trials discontinued (see below) Compared to many natural products, the structure of this substituted imidazole is simple. Girolline was reported to inhibit protein synthesis in eukaryotic target cells. The compound was of interest because it inhibited protein synthesis at the termination steps of the process rather than at the initiation or chain elongation steps like other known inhibitor compounds (e.g., emitine, homoharringtonine, anguidine). Although Girolline was moved through to Phase I clinical trials, the trials were stopped when some patients receiving the drug demonstrated significant hypertension as a side-effect.

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The simple structure of the compound facilitated the production of a modified synthetic analog, in the hope that the negative side effects could be eliminated. The resulting synthetic compound was shown not to have the bioactivity of the natural product, however. DIDEMNIN B Molecular Weight: 1112.35 g/mol Molecular Formula: C57H89N7O15 Source: The tunicate Trididemnum solidum (CHORDATA) Activity: Anti-cancer agent via protein synthesis inhibition Status: Withdrawn from clinical trials (see below) Didemnin B was originally isolated from the Caribbean tunicate Trididemnum solidum and first reported in the literature in 1981. Early investigation into the bioactivity of this compound revealed marked antiviral and cytotoxic activity in in vitro tests using standard mouse leukemia cell lines. Mechanistically, Didemnin B interrupts protein synthesis in target cells by binding non-competitively to palmitoyl protein thioesterase. Didemnin B was the first defined marine natural product to enter clinical trials as a potential anti-cancer drug. It proceeded through Phase I clinical trials as a prospective anticancer agent and entered into Phase II trials. Although the compound showed promising antitumor, antiviral and immunosuppressive activity, it also exhibited high toxicity, poor solubility, and a short bioactive lifespan. NCI withdrew the drug from clinical trials in the mid-1990s. Aplidine (Dehydrodidemnin B), a closely related natural product isolated from a different tunicate species, is currently in clinical trials as a potential anti-cancer drug. Although Didemnin B was never carried into Phase III trials, activity focused on developing the compound as a potential cancer treatment helped pave the way for the rest of the marine-derived products following it into the development pipeline. CURACIN A Molecular Weight: 373.596 g/mol Molecular Formula: C23H35NOS Source: Lyngbya majuscula (CYANOPHYTA) Activity: tubulin interactive compound Status: Preclinical First isolated from Lyngbya majuscula by Gerwick et al. and reported in 1994, this natural product appeared to be a very potent tubulin interactive compound. It proved to

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be highly insoluble, however, so much so that bioactivity could not be demonstrated with in vivo animal models. As a way around the insolubility issues presented by the natural curacin A, Wipf and colleagues developed a series of more soluble semi-synthetic variants using techniques of combinatorial chemistry to modify the basic structure. These compounds are currently undergoing preclinical evaluation as potential future drugs. DIAZONAMIDE A Molecular Weight: 765.64 g/mol Molecular Formula: C40H34Cl2N6O6 Source: The tunicate Diazona angulata (CHORDATA) Activity: Tubulin interactive agent Status: Preclinical The marine natural product Diazonamide A was first reported in 1991. It was extracted from the Philippine ascidian Diazona angulata by the William Fenical Chemistry Lab at the Scripp's Institution of Oceanography. In preliminary bioactivity screens the new compound killed lab-cultured colon cancer cells. More detailed characterization and biomedical evaluation was hindered by a lack of source material and failed efforts to locate and collect more of the rare species from which the compound was isolated. The structural complexity of the molecule made laboratory synthesis difficult, but funding from organizations like the American Cancer Society and the efforts of more than a dozen organic chemistry labs working toward the goal ultimately proved fruitful. In the process, synthetic chemists discovered that the original structure reported for the natural product was incorrect. When they finally synthesized an analog and then compared it to the original marine-derived product using X-ray crystalography and NMR, the differences became apparent. Subsequent to this discovery, a synthetic molecule structurally identical to natural diazonamide A has also been produced. Both analogs possess potent microtubulin interactive activity.

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Diazonamide A is an inhibitor of microtubule assembly, arresting the process of cell division in cultures exposed to treatment. Examination of treated cells reveals a loss of spindle microtubule assemblies and also microtubules associated with the interphase stage of the cell cycle. The precise mechanism of action is still under investiggation, but it has been postulated that the drug may bind tubulin at a unique site (e.g., distinct from dolastatin 10, vinca alkaloid, and other known binding sites), or that it may bind only weakly to unpolymerized tubulin but very strongly to fully formed microtubules. If the latter hypothesis is correct, it suggests that diazonamide A (and synthesized variants) could hold promise as a chemotherapeutant with unique microtubule inhibiting ability. DICTYOSTATIN Source: Unidentified Jamaican sponge of Family Corallistidae (PORIFERA) Activity: tubulin interactive compound. Status: Preclinical. Dictyostatin was first isolated in 1994 by Bob Pettit and his colleagues at Arizona State University from a sample of an unidntified dictyoceratid sponge of genus Spongia collected in Jamaica. The compound was later isolated from from a lithistid sponge of the family Corallistidae and has been more fully repurified and characterized by researchers at Harbor Branch Oceanographic Institution and elsewhere. Dictyostatin inhibits the growth of human cancer cells and has been shown active against certain Taxol-resistant tumors. Its mechanism of action appears to be prevention of the breakdown of tubulin during mitosis in a fashion similar to the successful cancer drug Taxol. Ian Patterson at Cambridge, in collaboration with Harbor Branch, has developed a synthesis method for dictyostatin, a 22-membered lactone with 11 stereocenters. In otther recent research, dictyostatin has also been syntthetically hybridized with the compound discodermolide. DISCODERMOLIDE Molecular Weight: 593.792 g/mol Molecular Formula: C33H55NO8 Source: The Caribbean deep-sea sponge Discodermia dissoluta (PORIFERA) Activity: tubule interactive agent Status: Phase I clinical trials Discodermolide, isolated from the Bahamian deep-sea sponge Discodermia dissoluta, is a promising marine-derived candidate for treating certain cancers. It was discovered in

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1987 by scientists with the Harbor Branch Division of Biomedical Marine Research. The drug, a macrolide (polyhydroxylated lactone), is a member of a structurallu diverse class of compounds called polyketides. It has a noteworthy chemical mechanism of action. It stabilizes the microtubules of target cells, essentially arresting them at a specific stage in the cell cycle and halting cell division. In addition to anticancer properties, discodermolide possesses immunosuppresive and cytotoxic activity. The pharmaceutical company Novartis Pharma AG licensed discodermolide for commercial development in 1998. The drug is currently in Phase I human clinical trials and continues to show promise in combating pancreatic cancer and many other drug-resistant cancers. Recently published reports offer up the exciting finding that combination drug treatment using discodermolide and Taxolآ in lung cancer patients exhibits several times the tumor fighting efficacy of either drug administered on its own. A number of synthetic discodermolide schemes have been elucidated, but so far none appear to be practical on a production-scale level. Biotech company Kosan Biosciences has used recombinant culture strategies to produce several novel discodermolide analogs currently under preclinical investigation. DOLASTATINS Molecular Weight (Dolastatin 10): 785.092 g/mol. Molecular Formula (Dolastatin 10): C42H68N6O6S. Source: Indian Ocean sea hare Dollabella auricularia (MOLLUSCA) Activity: Tubulin interactive agents Status: Phase II, Phase I clinical trials. Dolastatin 10 and dolastatin 15 were isolated from the Indian Ocean sea hare Dollabella auricularia. These small linear peptide molecules are promising anti-cancer drugs showing potency against breast and liver cancers, solid tumors and some leukemias. Preclinical research indicated potency in experimental antineoplastic and tubulin assembly systems. These products had been hypothesized as being microbial products and not actually produced by

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D. auricularia. The peptides comprising the dolastatins contained an assortment of amino acids that pointed to a probable cynobacterial (blue-green "algae") origin. Later research has shown this to be the case, and both dolastatin 10 and the structurally similar compound Simplostatin 1 have both been isolated from marine cyanobacteria. Rather than being a microbial symbiont, the cyanobacterium that produced dolastatin 10 is a mat-forming species known to be grazed by D. auricularia. The dolastatins are mitotic inhibitors. They interfere with tubulin formation and thereby disrupt cell division by mitosis. These molecules bind to tubulin at a location known as the vinca/peptide region. This region is the target for several other structurally complex natural products, including the sponge product hemiasterlin. Phase II clinical trials of dolastatin 10 in patients with indolent lymphoma, Waldenstrom's macroglobulinemia, and chronic lymphocytic leukemia (CLL) have been completed. Performance against solid tumors was lacking, but dolastatin 10 remains a good candidate for combination drug regimes that also include vinca alkaloids or possibly bryostatin 1. The development of synthetic analogs to the dolastatins has been vigorously pursued. Several such analogs have been successfully synthesized and some of these are now also in the pre-clinical/clinical pipeline. ILX651 and LU103793 are two such compounds. LU103793 has been evaluated in a number of Phase I trials. ILX651 is currently being assessed as a treatment for several tumor types including colorectal, lung, melanoma, renal, and pancreatic cancers. ELEUTHEROBIN Source: The octocorals Eleutherobia sp., Erythropodium caribaeorum (CNIDARIA) Activity: Tubulin interactive agent Status: Preclinical Eleutherobin was first found in extracts made from the octocoral Eleutherobia sp. collected in Australia. The species of Eleutherobia was first described by Scripps Institution of Oceanography's Bill Fenical in 1993. Eleutherobin was extracted from the coral two years later by Fenical's post-doctoral student Thomas Lindel. The rarity of Eleutherobia sp. greatly impeded development since it was the only known source of the drug. More recently, however, eleutherobin was isolated from the Caribbean encrusting coral Erythropodium caribaeorum by the research group led by Ray Andersen of the University of British Columbia. The compound has also been recovered from aquacultured E. caribaeorum. Currently synthetic production methods are being explored.

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The reported bioactivity of this natural product, currently under preclinical investigation, is as a microtubule binding agent similar to the anti-cancer drug taxol. There is hope that eleutherobin will eventually prove similarly effective but with perhaps fewer side effects (e.g., immune system suppression, nausea, hair loss) than taxol. HALICHONDRINS Molecular Weight (Halichondrin B): 1111.31 g/mol Molecular Formula (Halichondrin B): C60H86O19 Source: The Japanese sponge Halichondria okadai (PORIFERA) Activity: tubulin interactive agent Status: Synthetic analogs are currently in clinical trials. A compound given the name halichondrin B, belonging to a chemical family known as the macrolides, was isolated by Uemura et al (1985) from the Japanese sponge Halichondria okadai. Since this time, several similar macrolides have been found in an array of Pacific and Indian Ocean sponge genera including Axinella, Phakellia, and Lissodendroryx. Initial investigations into the bioactivity of the compound revealed that halichondrin B apparently bound tubilin at a site close to the so-called vinca site and altered tubulin depolymerization. Early experimental work also demonstrated in vivo activity, but further work advancing the compound was hindered by lack of the natural product. The National Cancer Institutes provided funds for a New Zealand research consortium to trawl-harvest the one metric ton of the deepwater sponge Lissodendoryx sp. that would be needed to yield sufficient halichondrins to continue evaluation and development. NCI also funded successful efforts to culture this sponge in shallow water. At approximately the same time, a Harvard synthetic chemistry lab succeeded in the total synthesis of halchondrin B and also the related structure norhalichondrin B. This group produced supplies of the compound via synthesis for use in NCI-sponsored clinical trials. This synthetic work was expanded upon by scientists working for Japanese pharmaceutical company Eisai, who successfully produced a range of halichondrin B variants that remained bioactive but were also more structurally stable than their natural counterpart. One of the Eisai halichondrin analogs, E7389, is currently in phase I clinical studies under the auspices of NCI. HEMIASTERLINS Molecular Weight (Hemiasterlin): 526.711 g/mol. Molecular Formula (Hemiasterlin): C30H46N4O4.

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Source: Sponges of genus Auletta, Siphonochalin (PORIFERA). Activity: cytotoxic and tubulin interactive agents. Status: Preclinical This class of novel marine oligopeptides have been shown to be potent antitumor agents. Representatives of this class of natural products have been isolated from extracts prepared from sponges residing in two distinct genera (Auletta; Siphonochalina). Three different hemiasterlins with drug development potential (hemiasterlin, hemiasterlin A, hemiasterlin C) have been the subject of chemical and biological investigations. These molecules exhibit cytotoxic and antitubulin activity similar to that seen in the dolastatins. Mitotic inhibition occurs through binding to tubulin at the vinca/peptide region in a manner similar to dolastatin. The production of synthetic hemiasterlin analogs, facilitated by the simple tripeptide molecular structure, has been achieved. Studies have demonstrated that one analog, HTI-286, effectively inhibits tubulin polymerization, disrupts cell microtubule organization, and induces mitotic arrest and apoptosis. LATRUNCULINS Molecular Weight (Latrunculin A): 421.551 g/mol. Molecular Formula (Latrunculin A): C22H31O5S. Source: The Red Sea sponge Latrunculia magnifica (PORIFERA). Activity: Actin interactive agent. Status: Preclinical, see below. The Latrunculins were first isolated from the Red Sea sponge Latrunculia magnifica. Latrunculins have since been found in the sponge Negombata magnifica. Structural similarity of these compounds with those produced via polyketide synthesis in microbes suggests the source of the latrunculins is actually a member of the sponge-associated microbial community. Two compounds tested, latrunculins A and B, have been shown to be cytotoxic, and the toxicity appears related to the compounds' ability to disrupt actin polymerization,

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microfilament organization, etc. Cell shape, cytokinesis, and microfilament-mediated processes such as fertilization and early development are altered through exposure to the drugs. Although interest in this and other actin interative agents remains high, a general lack of significant in vivo activity in preclinical evaluations has hindered drug develpment. If novel targeted strategies for drug delivery like the use of monoclonal antibodies or peptides as carriers can be developed, the potential of the latrunculins and similar natural products as drug candidates may improve. LAULIMALIDE (and Synthetic Analogs) Source: The Pacific sponge Cacospongia mycofijiensis; also Hyatella sp., Fasciospongia sp., and Dactylospongia sp. (PORIFERA) Activity: tubulin interactive agent. Status: Preclinical. Sometimes referred to as the fijianolides, the natural products laulimide and (the significantly less bioactive) isolaulimide were first extracted from the Pacific sponge Cacospongia mycofijiensis. Since their initial discovery, the compounds have been found in other unrelated sponge genera, and also in extracts from a sponge-grazing chromodorid nudibranch. A number of laboratory synthetic pathways for producing analogs of these natural compounds have been published. The bioactivity displayed by these compounds is as microtubule stabilizing agents potentially arresting the development of target cells. The exact mode of action with regard to how and where the molecules bind tubulin has not been reported, but the activity profiles are clearly different from that of other microtubule-binding agents such as paclitaxel. While naturally occurring laulimalide shows effectiveness against paclitaxel-resistant cells, the compound is intrinsically unstable. Recently, Mooberry and colleagues (2004) synthesized several laulimalide analogs designed to address this stability issue. Some of these retain the unique biological activities of the natural molecule, including effectiveness against paclitaxel- and epothilone-resistant cell lines. PELORUSIDE A Source: The New Zealand sponge Mycale hentscheli (PORIFERA). Activity: tubulin interactive agent. Status: Preclinical University of Victoria scientist Peter Northcote and colleagues recovered the compound Peloruside A from the sponge Mycale hentscheli collected from Pelorus Sound, New Zealand. Not much has been published on this tubulin interactive compound since the

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initial report in 2000. But, this relatively simple compound may facilitate exploration of synthetic and semi-synthetic derivatives if the basic bioactivity looks promising. Dr Northcote reports that Peloruside A shares a molecular target with the popular commercial cancer therapy agent paclitaxel. Peloruside A appears to bind tubulin and arrests target cell development at the G2-M transition stage of the cell cycle, triggering apoptosis ('cell suicide') before mitosis (M Phase) can begin. Very recently, the compound was licensed from the University of Texas Southwestern Medical Center at Dallas and Victoria University of Wellington, New Zealand, by biopharmaceutical company Reata Pharmaceuticals, Inc.. Scientists from this company recently completed the first total synthesis of peloruside A, and they suggest the compound has a chemical scaffold that is amenable to modification for the purpose of drug development. They believe Peloruside A could enter clinical trials as early as 2007. Efforts are also ongoing by researchers from New Zealand's National Institute of Water and Atmospheric Research (NIWA) to successfully culture the sponge while retaining bioactivity similar to that found in wild stocks. SARCODICTYINS Molecular Weight (Sarcodictyin A): 496.595 g/mol. Molecular Formula (Sarcodictyin A): C28H36N2O6. Source: The corals Sarcodictyon roseum, Eleutherobia aurea, and others (CNIDARIA) Activity: Tubulin interactive agent Status: Preclinical In the mid 1980s, natural products called sarcodictyins were isolated from two Mediterranean coral species, Sarcodictyon roseum and Eleutherobia aurea. Although the were shown to have an interesting structural configuration, initial screens determined that these compounds lacked discernable bioactivity. This assessment was set aside with the 1997 discovery of sarcodictyin activity as tubulin interactive agents. In 2003, A new cytotoxic diterpene, (Z)-sarcodictyin A, was reported isolated from the Japanese soft coral Bellonella albiflora.

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Preclinical research focusing on the sarcodictyins is being conducted by drug company Pharmacia-Upjohn, who were first to report tubulin binding by these compounds. These compounds have also been the subject of intriguing combinatorial synthetic research in which novel hybrid molecules were produced by combining the base structures of sarcodictyins with those of another cnidarian-derived class of tubulin interactive natural products, the eleutherobins. VITILEVUAMIDE Source: The the ascidians Didemnum cuculiferum, Polysyncraton lithostrotum (CHORDATA) Activity: Tubulin interactive agent. Status: Preclinical Vitilevuamide is a bioactive cyclic peptide has been isolated from the ascidians Didemnum cuculiferum and Polysyncraton lithostrotum (the same animal is the source of the antimicrobial/antitumor compound namenamicin). Vitilevuamide is one of several novel tubulin interactive agents recently discovered from marine invertebrate sources. Research on the mechanism of action of this two-ringed marine peptide reveal that vitilevuamide inhibit tubulin polymerization and can arrest the cell cycle of target cells in the G2/M phase. The drug exhibits activity in vivo against the P388 lymphocytic leukemia line. An intriguing finding is that tubulin binding and inhibition by this compound occurs at a site on the tubulin molecule distinct from the interaction sites of dolastatin 10, colchicine, and the vinca alkaloids. Vitilevuamide is currently in the preclinical evaluation phase as a potential anti-cancer agent.

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SALICYLIHALAMIDES Molecular Weight (Salicylihalamide A): 439.544 g/mol. Molecular Formula(Salicylihalamide A): C26H33NO5. Source: The sponge Haliclona sp. (PORIFERA) Activity: Vo-ATPase inhibitor Status: Preclinical The Vo-ATPases are a group of eukaryotic enzymes whose principal role is to pump hydrogen ions across cell vacuolar membranes. Salicylihalamides A and B, isolated from the sponge Haliclona sp. collected off of Western Australia, displays Vo-ATPase inhibitory activity that may eventually be incorporated into a drug designed to target these cell components. In addition to having potential as anti-tumor compounds, Vo-ATPases like the salicylihalamides may be capable of mediating the process of bone resorption. In this capacity, the compounds might form the basis for treatment of osteoporosis and similar diseases.

References

1. Barsanti, L. and P. Gualtieri, Algae Anatomy, Biochemistry, and Biotechnology. 2006, Boca Raton London New York: Taylor & Francis Group. 301.

2. Richmond, A., Handbook of Microalgal Culture: Biotechnology and Applied Phycology. 2004: Blackwell Science Ltd. 566.

3. Andersen, R.A., Algal Culturing Techniques, ed. F. Cynar. 2005, California: Elsevier Academic Press. 578.

4. Lekang, O.-I., Aquaculture Engineering. 2007, UK: Blackwell Publishing Ltd. 340.

5. Railkin, A.I., MARINE BIOFOULING Colonization Processes and Defenses, ed. C.P. LLC. 2004, Boca Raton London New York Washington, D.C. 303.

6. Matsunaga, T., et al., Marine Biotechnology I. 2005, Berlin Heidelberg: Springer-Verlag 262.

7. Dunham, R.A., Aquaculture and Fisheries Biotechnology. 2004, Alabama USA: CABI Publishing. 372.

8. Bhakuni, D.S. and D.S. Rawat, Bioactive Marine Natural Products. 2005, New York, USA: Springer. 382.

9. Beaumont, A.R. and K. Hoare, Biotechnology and Genetics in Fisheries and Aquaculture. 2003 USA: Blackwell Science Ltd. 158.

10. Lutz, C.G., Practical Genetics for Aquaculture. 2001: Blackwell Science Ltd. 235.

11. ENVIRONMENTAL RISK ASSESSMENT OF GENETICALLY MODIFIED ORGANISMS, ed. A.R. Kapuscinski, et al. Vol. 3. Methodologies for Transgenic Fish. 2007: CABI.

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