PEGylated Recombinant Aplysia punctata Ink Toxin Depletes Arginine and Lysine and Inhibits the Growth of Tumor Xenografts.

In recent years, a novel treatment method for cancer has emerged, which is based on the starvation of tumors of amino acids like arginine. The deprivation of arginine in serum is based on enzymatic degradation and can be realized by arginine deaminases like the l-amino acid oxidase found in the ink toxin of the sea hare Aplysia punctata. Previously isolated from the ink, the l-amino acid oxidase was described to oxidate the essential amino acids l-lysine and l-arginine to their corresponding deaminated alpha-keto acids. Here, we present the recombinant production and functionalization of the amino acid oxidase Aplysia punctata ink toxin (APIT). PEGylated APIT (APIT-PEG) increased the blood circulation time. APIT-PEG treatment of patient-derived xenografted mice shows a significant dose-dependent reduction of tumor growth over time mediated by amino acid starvation of the tumor. Treatment of mice with APIT-PEG, which led to deprivation of arginine, was well tolerated.

Current Trends and New Challenges in Marine Phycotoxins.

Marine phycotoxins are a multiplicity of bioactive compounds which are produced by microalgae and bioaccumulate in the marine food web. Phycotoxins affect the ecosystem, pose a threat to human health, and have important economic effects on aquaculture and tourism worldwide. However, human health and food safety have been the primary concerns when considering the impacts of phycotoxins. Phycotoxins toxicity information, often used to set regulatory limits for these toxins in shellfish, lacks traceability of toxicity values highlighting the need for predefined toxicological criteria. Toxicity data together with adequate detection methods for monitoring procedures are crucial to protect human health. However, despite technological advances, there are still methodological uncertainties and high demand for universal phycotoxin detectors. This review focuses on these topics, including uncertainties of climate change, providing an overview of the current information as well as future perspectives.

Voltage-Gated Sodium Channels: A Prominent Target of Marine Toxins.

Voltage-gated sodium channels (VGSCs) are considered to be one of the most important ion channels given their remarkable physiological role. VGSCs constitute a family of large transmembrane proteins that allow transmission, generation, and propagation of action potentials. This occurs by conducting Na+ ions through the membrane, supporting cell excitability and communication signals in various systems. As a result, a wide range of coordination and physiological functions, from locomotion to cognition, can be accomplished. Drugs that target and alter the molecular mechanism of VGSCs' function have highly contributed to the discovery and perception of the function and the structure of this channel. Among those drugs are various marine toxins produced by harmful microorganisms or venomous animals. These toxins have played a key role in understanding the mode of action of VGSCs and in mapping their various allosteric binding sites. Furthermore, marine toxins appear to be an emerging source of therapeutic tools that can relieve pain or treat VGSC-related human channelopathies. Several studies documented the effect of marine toxins on VGSCs as well as their pharmaceutical applications, but none of them underlined the principal marine toxins and their effect on VGSCs. Therefore, this review aims to highlight the neurotoxins produced by marine animals such as pufferfish, shellfish, sea anemone, and cone snail that are active on VGSCs and discuss their pharmaceutical values.

Specific Antiproliferative Properties of Proteinaceous Toxin Secretions from the Marine Annelid Eulalia sp. onto Ovarian Cancer Cells.

As Yondelis joins the ranks of approved anti-cancer drugs, the benefit from exploring the oceans' biodiversity becomes clear. From marine toxins, relevant bioproducts can be obtained due to their potential to interfere with specific pathways. We explored the cytotoxicity of toxin-bearing secretions of the polychaete Eulalia onto a battery of normal and cancer human cell lines and discovered that the cocktail of proteins is more toxic towards an ovarian cancer cell line (A2780). The secretions' main proteins were identified by proteomics and transcriptomics: 14-3-3 protein, Hsp70, Rab3, Arylsulfatase B and serine protease, the latter two being known toxins. This mixture of toxins induces cell-cycle arrest at G2/M phase after 3h exposure in A2780 cells and extrinsic programmed cell death. These findings indicate that partial re-activation of the G2/M checkpoint, which is inactivated in many cancer cells, can be partly reversed by the toxic mixture. Protein-protein interaction networks partake in two cytotoxic effects: cell-cycle arrest with a link to RAB3C and RAF1; and lytic activity of arylsulfatases. The discovery of both mechanisms indicates that venomous mixtures may affect proliferating cells in a specific manner, highlighting the cocktails' potential in the fine-tuning of anti-cancer therapeutics targeting cell cycle and protein homeostasis.

Marine Compounds and Cancer: Updates 2020.

By the end of the year 2020, there are nine marine-derived anticancer drugs available on the market, and the field is currently growing exponentially [...].

Marine Toxins Targeting Kv1 Channels: Pharmacological Tools and Therapeutic Scaffolds.

Toxins from marine animals provide molecular tools for the study of many ion channels, including mammalian voltage-gated potassium channels of the Kv1 family. Selectivity profiling and molecular investigation of these toxins have contributed to the development of novel drug leads with therapeutic potential for the treatment of ion channel-related diseases or channelopathies. Here, we review specific peptide and small-molecule marine toxins modulating Kv1 channels and thus cover recent findings of bioactives found in the venoms of marine Gastropod (cone snails), Cnidarian (sea anemones), and small compounds from cyanobacteria. Furthermore, we discuss pivotal advancements at exploiting the interaction of κM-conotoxin RIIIJ and heteromeric Kv1.1/1.2 channels as prevalent neuronal Kv complex. RIIIJ's exquisite Kv1 subtype selectivity underpins a novel and facile functional classification of large-diameter dorsal root ganglion neurons. The vast potential of marine toxins warrants further collaborative efforts and high-throughput approaches aimed at the discovery and profiling of Kv1-targeted bioactives, which will greatly accelerate the development of a thorough molecular toolbox and much-needed therapeutics.

Marine Toxins and Nociception: Potential Therapeutic Use in the Treatment of Visceral Pain Associated with Gastrointestinal Disorders.

Visceral pain, of which the pathogenic basis is currently largely unknown, is a hallmark symptom of both functional disorders, such as irritable bowel syndrome, and inflammatory bowel disease. Intrinsic sensory neurons in the enteric nervous system and afferent sensory neurons of the dorsal root ganglia, connecting with the central nervous system, represent the primary neuronal pathways transducing gut visceral pain. Current pharmacological therapies have several limitations, owing to their partial efficacy and the generation of severe adverse effects. Numerous cellular targets of visceral nociception have been recognized, including, among others, channels (i.e., voltage-gated sodium channels, VGSCs, voltage-gated calcium channels, VGCCs, Transient Receptor Potential, TRP, and Acid-sensing ion channels, ASICs) and neurotransmitter pathways (i.e., GABAergic pathways), which represent attractive targets for the discovery of novel drugs. Natural biologically active compounds, such as marine toxins, able to bind with high affinity and selectivity to different visceral pain molecular mediators, may represent a useful tool (1) to improve our knowledge of the physiological and pathological relevance of each nociceptive target, and (2) to discover therapeutically valuable molecules. In this review we report the most recent literature describing the effects of marine toxin on gastrointestinal visceral pain pathways and the possible clinical implications in the treatment of chronic pain associated with gut diseases.

The "Utility" of Highly Toxic Marine-Sourced Compounds.

Currently a few compounds isolated from marine sources have become drugs, mainly directed towards cancer and pain. Compounds from marine sources have exquisite potencies against eukaryotic cells, as they act as protective agents against attack by predators in the marine environment. Their toxicities act as a "double-edged sword" as they are often too toxic for direct use in humans and thus have to be chemically modified. By linking suitably modified compounds to monoclonal antibodies directed against specific epitopes in mammalian cancer cells, they can be delivered to a specific cell type in humans. This review updates and extends an article published in early 2017, demonstrating how by careful chemical modifications, highly toxic compounds, frequently peptidic in nature, can be utilized as antitumor drug candidates. The antibody-drug- conjugates (ADCs) discussed are those that are currently in clinical trials listed in the NIH Clinical Trials Registry as, "currently active, recruiting or in some cases, recently completed". There are also some ADCs discussed that are at the advanced preclinical stage, that in some cases, are repurposing current drug entities, and the review finishes with a short discussion of the aplyronines as potential candidate warheads as a result of scalable synthetic processes.

The Marine Dinoflagellate Alexandrium minutum Activates a Mitophagic Pathway in Human Lung Cancer Cells.

Marine dinoflagellates are a valuable source of bioactive molecules. Many species produce cytotoxic compounds and some of these compounds have also been investigated for their anticancer potential. Here, we report the first investigation of the toxic dinoflagellate Alexandrium minutum as source of water-soluble compounds with antiproliferative activity against human lung cancer cells. A multi-step enrichment of the phenol⁻water extract yielded a bioactive fraction with specific antiproliferative effect (IC50 = 0.4 µg·mL-1) against the human lung adenocarcinoma cells (A549 cell line). Preliminary characterization of this material suggested the presence of glycoprotein with molecular weight above 20 kDa. Interestingly, this fraction did not exhibit any cytotoxicity against human normal lung fibroblasts (WI38). Differential gene expression analysis in A549 cancer cells suggested that the active fraction induces specific cell death, triggered by mitochondrial autophagy (mitophagy). In agreement with the cell viability results, gene expression data also showed that no mitophagic event was activated in normal cells WI38.

Ascidian Toxins with Potential for Drug Development.

Ascidians (tunicates) are invertebrate chordates, and prolific producers of a wide variety of biologically active secondary metabolites from cyclic peptides to aromatic alkaloids. Several of these compounds have properties which make them candidates for potential new drugs to treat diseases such as cancer. Many of these natural products are not produced by the ascidians themselves, rather by their associated symbionts. This review will focus mainly on the mechanism of action of important classes of cytotoxic molecules isolated from ascidians. These toxins affect DNA transcription, protein translation, drug efflux pumps, signaling pathways and the cytoskeleton. Two ascidian compounds have already found applications in the treatment of cancer and others are being investigated for their potential in cancer, neurodegenerative and other diseases.

The anti-tumour activity of rLj-RGD4, an RGD toxin protein from Lampetra japonica, on human laryngeal squamous carcinoma Hep-2 cells in nude mice.

PURPOSE: The objective of this study is to investigate the antiproliferative activity and mechanism of integrin-binding rLj-RGD4 in a Hep-2 human laryngeal carcinoma-bearing nude mouse model. METHODS: Human laryngeal squamous carcinoma cells (Hep-2) were inoculated subcutaneously into the axilla of nude mice to generate a Hep-2 human laryngeal carcinoma-bearing nude mouse model. When the Hep-2 xenograft model was successfully established, the animals were randomly separated into five groups. Three groups were treated with different dosages of rLj-RGD4. Cisplatin was administered to the positive control group, and normal saline (NaCl) was administered to the negative control group for 3 weeks. The body weights and the survival of the nude mice were evaluated, and the volumes and weights of the solid tumours were measured. The mechanism underlying rLj-RGD4 inhibition of tumour growth in transplanted Hep-2 human laryngeal carcinoma-bearing nude mice was evaluated by haematoxylin-eosin (HE) staining, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL), measurement of intratumoural microvessel density (MVD), Western blotting, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: The tumour volumes and weights of the treatment groups were reduced compared with the model group, and survival times were improved by rLj-RGD4 treatment in Hep-2 human laryngeal carcinoma-bearing nude mice. The number of apoptotic Hep-2 human cells and intratumoural MVD significantly decreased after the administration of rLj-RGD4. In the xenograft tissue of animals treated with rLj-RGD4, FAK, PI3K, and Akt expression was unaltered, whereas P-FAK, P-PI3K, Bcl-2, P-Akt, and VEGF levels were down-regulated. In addition, activated caspase-3, activated caspase-9, and Bax levels were up-regulated. CONCLUSION: rLj-RGD4 exhibits potent in vivo activity and inhibits the growth of transplanted Hep-2 human laryngeal carcinoma cells in a nude mouse model. Thus, these results indicate that the recombinant RGD toxin protein rLj-RGD4 may serve as a potent clinical therapy for human laryngeal squamous carcinoma.

Augmenting anti-cancer natural products with a small molecule adjuvant.

Aquatic microbes produce diverse secondary metabolites with interesting biological activities. Cytotoxic metabolites have the potential to become lead compounds or drugs for cancer treatment. Many cytotoxic compounds, however, show undesirable toxicity at higher concentrations. Such undesirable activity may be reduced or eliminated by using lower doses of the cytotoxic compound in combination with another compound that modulates its activity. Here, we have examined the cytotoxicity of four microbial metabolites [ethyl N-(2-phenethyl) carbamate (NP-1), Euglenophycin, Anabaenopeptin, and Glycolipid 652] using three in vitro cell lines [human breast cancer cells (MCF-7), mouse neuroblastoma cells (N2a), and rat pituitary epithelial cells (GH4C1)]. The compounds showed variable cytotoxicity, with Euglenophycin displaying specificity for N2a cells. We have also examined the modulatory power of NP-1 on the cytotoxicity of the other three compounds and found that at a permissible concentration (125 µg/mL), NP-1 sensitized N2a and MCF-7 cells to Euglenophycin and Glycolipid 652 induced cytotoxicity.

Drug development for pediatric neurogenic bladder dysfunction: dosing, endpoints, and study design.

Pediatric drug development is challenging when a product is studied for a pediatric disease that has a different underlying etiology and pathophysiology compared to the adult disease. Neurogenic bladder dysfunction (NBD) is such a therapeutic area with multiple unsuccessful development programs. The objective of this study was to critically evaluate clinical trial design elements that may have contributed to unsuccessful drug development programs for pediatric NBD. Trial design elements of drugs tested for pediatric NBD were identified from trials submitted to the U.S. Food and Drug Administration. Data were extracted from publically available FDA reviews and labeling and included trial design, primary endpoints, enrollment eligibilities, and pharmacokinetic data. A total of four products were identified. Although all four programs potentially provided clinically useful information, only one drug (oxybutynin) demonstrated efficacy in children with NBD. The lack of demonstrable efficacy for the remainder of the products illustrates that future trials should give careful attention to testing a range of doses, using objectively measured, clinically meaningful endpoints, and selecting clinical trial designs that are both interpretable and feasible. Compiling the drug development experience with pediatric NBD will facilitate an improved approach for future drug development for this, and perhaps other, therapeutic areas.

Sea-originated cytotoxic substances.

Cancer accounted for huge number of deaths, which represents about 13% of all deaths worldwide, and the number of the deaths due to cancer is increasing. Natural products and their synthetic analogs are widely used as antitumor drugs. As represented by these drugs, many anticancer drugs originated from cytotoxic compounds. Marine natural products are a gold mine of strong bioactive compounds with unique structures created in evolution of organisms over hundred million years. However, in the field of drug discovery, most studies have focused on plant essences and bacterial metabolites, and candidate compounds from marine origin are still remaining relatively unexplored.

Natural toxins and their therapeutic potential.

Plants have been extensively investigated for exploring their therapeutic potentials, but there are comparatively scanty reports on drugs derived from animal kingdom, except for hormones. During last decade, the toxins that are used for defense by the animals, have been isolated and found useful tools for physiological and pharmacological studies, besides giving valuable leads to drug development. Toxins with interesting results have been isolated from the venoms of snakes, scorpions, spiders, snails, lizards, frogs and fish. The present review describe about some toxins as drugs and their biological activities. Some fungal, bacterial and marine toxins have also been covered in this article.

Isolation and cDNA cloning of type 2 sodium channel peptide toxins from three species of sea anemones (Cryptodendrum adhaesivum, Heterodactyla hemprichii and Thalassianthus aster) belonging to the family Thalassianthidae.

The crude extracts from three species of sea anemones (Cryptodendrum adhaesivum, Heterodactyla hemprichii and Thalassianthus aster) belonging to the family Thalassianthidae exhibited potent lethality to freshwater crabs (Potamon dehaani). Regardless of the species, high and low molecular weight toxins were found in gel filtration of the crude extract. Following reverse-phase HPLC of the low molecular weight toxin fractions, one toxin (δ-TLTX-Ca1a), two toxins (δ-TLTX-Hh1a and c) and one toxin (δ-TLTX-Ta1a) were isolated from C. adhaesivum, H. hemprichii and T. aster, respectively. Based on the determined N-terminal amino acid sequences, the cDNAs encoding δ-TLTX-Ca1a, δ-TLTX-Hh1x (not assignable to either δ-TLTX-Hh1a or δ-TLTX-Hh1c) and δ-TLTX-Ta1a were successfully cloned by both 3' and 5' RACE methods. In common with the three toxins, the precursor is composed of a signal peptide (19 amino acid residues), propart (16 residues) and mature portion (49 residues), similar to those of many sea anemone peptide toxins. The deduced amino acid sequences showed that the three toxins are closely similar to one another, being all new members of the type 2 sea anemone sodium channel peptide toxin family.

Marine natural product drug discovery: Leads for treatment of inflammation, cancer, infections, and neurological disorders.

Natural products, secondary metabolites, isolated from plants, animals and microbes are important sources for bioactive molecules that in many cases have been developed into treatments for diseases. This review will focus on describing the potential for finding new treatments from marine natural products for inflammation, cancer, infections, and neurological disorders. Historically terrestrial natural products have been studied to a greater extent and such classic drugs as aspirin, vincristine and many of the antibiotics are derived from terrestrial natural products. The need for new therapeutics in the four areas mentioned is dire. Within the last 30 years marine natural products, with their unique structures and high level of halogenation, have shown many promising activities against the inflammatory response, cancer, infections and neurological disorders. The review will outline examples of such compounds and activities.

Reports of clinical benefit of plitidepsin (Aplidine), a new marine-derived anticancer agent, in patients with advanced medullary thyroid carcinoma.

OBJECTIVES: To assess clinical benefit of plitidepsin (Aplidine) in patients with advanced medullary thyroid carcinoma (MTC). MATERIALS AND METHODS: We retrospectively reported the outcome of 10 patients with advanced MTC among 215 patients who have entered the phase I program with plitidepsin. RESULTS: Median number of cycles was 5. Using World Health Organization criteria, 1 among 5 patients with measurable disease displayed a confirmed partial response, whereas 8 patients experienced a stable disease, and 1 patient had a progressive disease, corresponding to a disease control rate of 90%. Two patients treated at the maximum tolerated dose experienced muscular dose-limiting toxicity possibly related to palmitoyl transferase inhibition. One of these 2 patients was able to continue therapy with no dose reduction with the prophylactic addition of l-carnitine, which is used in the treatment of the carnitine palmitoyl transferase deficiency type 2. DISCUSSION: Plitidepsin seems to be able to induce clinical benefit in patients with pretreated MTC, and its toxicity has been manageable at the recommended dose.