Identification of a Double-ß-Defensin with Multiple Antimicrobial Activities in a Marine Invertebrate.

ß-Defensins are a family of cysteine-rich antimicrobial peptides that are generally monodomain. Interestingly, the avian ß-defensin 11 (AvBD11) is unique, with two ß-defensin motifs with a broad range of antimicrobial activities. However, a double-sized ß-defensin has not been identified and functionally characterized in invertebrates. In this study, we cloned and identified a double-ß-defensin in shrimp Litopenaeus vannamei (named LvDBD) and explored its potential roles during infection with shrimp pathogens Vibrio parahaemolyticus and white spot syndrome virus (WSSV). LvDBD is an atypical double-sized defensin, which is predicted to possess two motifs related to ß-defensin and six disulfide bridges. The RNA interference-mediated knockdown of LvDBD in vivo results in phenotypes with increased bacterial loads, rendering the shrimp more susceptible to V. parahaemolyticus infection, which could be rescued by the injection of recombinant LvDBD protein. In vitro, rLvDBD could destroy bacterial membranes and enhance hemocyte phagocytosis, possibly attributable to its affinity to the bacterial wall components LPS and peptidoglycan. In addition, LvDBD could interact with several viral envelope proteins to inhibit WSSV proliferation. Finally, the NF-κB transcription factors (Dorsal and Relish) participated in the regulation of LvDBD expression. Taken together, these results extend the functional understanding of a double-ß-defensin to an invertebrate and suggest that LvDBD may be an alternative agent for the prevention and treatment of diseases caused by V. parahaemolyticus and WSSV in shrimp.

Effects of copper on non-specific immunity and antioxidant in the oriental river prawn (Macrobrachium nipponense).

The copper, as heavy metal has important impacts on the antioxidant and immune defense systems in aquatic organisms, and the toxic effects of copper can be accumulated and magnified with the food chain, thus posing a threat to food safety as well as ecosystems. This study explored the response of the antioxidant system and non-specific immunity in M. nipponense to copper stress. Low concentration of copper (0.05, 0.1 mg L-1) had positive effects on the non-specific immunity in M. nipponense, while the non-specific immunity in M. nipponense could be affect negatively or even be inhibited by high copper concentration (0.15 mg L-1). Even low concentrations of copper could cause oxidative stress, and high copper concentration (0.15 mg L-1) could induce oxidative damage and even apoptosis, and thus causing damage to the antioxidant defense system in M. nipponense. Low concentration of copper could affect the gill and hepatopancreas structure in M. nipponense, but high level oxidative stress caused by high copper concentration could cause oxidative damage to these tissue, resulting in the destruction of gill and hepatopancreas. This study provides the safety concentration for using copper-containing fish drugs in the actual culture of M. nipponense and provides basic data for the toxicity mechanism of copper to M. nipponense.

SpCrus2 Glycine-Rich Region Contributes Largely to the Antiviral Activity of the Whole-Protein Molecule by Interacting with VP26, a WSSV Structural Protein.

Crustins are cysteine-rich cationic antimicrobial peptides with diverse biological functions including antimicrobial and proteinase inhibitory activities in crustaceans. Although a few crustins reportedly respond to white spot syndrome virus (WSSV) infection, the detailed antiviral mechanisms of crustins remain largely unknown. Our previous research has shown that SpCrus2, from mud crab Scylla paramamosain, is a type II crustin containing a glycine-rich region (GRR) and a cysteine-rich region (CRR). In the present study, we found that SpCrus2 was upregulated in gills after WSSV challenge. Knockdown of SpCrus2 by injecting double-stranded RNA (dsSpCrus2) resulted in remarkably increased virus copies in mud crabs after infection with WSSV. These results suggested that SpCrus2 played a critical role in the antiviral immunity of mud crab. A GST pull-down assay showed that recombinant SpCrus2 interacted specifically with WSSV structural protein VP26, and this result was further confirmed by a co-immunoprecipitation assay with Drosophila S2 cells. As the signature sequence of type II crustin, SpCrus2 GRR is a glycine-rich cationic polypeptide with amphipathic properties. Our study demonstrated that the GRR and CRR of SpCrus2 exhibited binding activities to VP26, with the former displaying more potent binding ability than the latter. Interestingly, pre-incubating WSSV particles with recombinant SpCrus2 (rSpCrus2), rGRR, or rCRR inhibited virus proliferation in vivo; moreover, rSpCrus2 and rGRR possessed similar antiviral abilities, which were much stronger than those of rCRR. These findings indicated that SpCrus2 GRR contributed largely to the antiviral ability of SpCrus2, and that the stronger antiviral ability of GRR might result from its stronger binding activity to the viral structural protein. Overall, this study provided new insights into the antiviral mechanism of SpCrus2 and the development of new antiviral drugs.

Antarctic krill-derived peptides with consecutive Glu residues enhanced iron binding, solubility, and absorption.

Three peptides containing three glutamic acid (Glu) residues at different positions derived from Antarctic krill were obtained to investigate their iron-binding properties, digestive stability, and effectiveness on enhancing iron solubility and absorption. Results indicated that Fe2+ bound to the carbonyl, carboxyl, or hydroxyl groups of DELEDSLER, EEEFDATR, and DTDSEEEIR at stoichiometric ratios of 0.453, 0.466, and 0.490, respectively. DTDSEEEIR with three consecutive Glu in the middle of the sequence possessed higher iron-binding ability and iron release potential than EEEFDATR with three consecutive Glu in the N-terminal, and DELEDSLER with three discontinuous Glu showed the lowest values. Although EEEFDATR showed remarkably lower digestion stability than DTDSEEEIR, the effect of EEEFDATR-iron on iron solubility and absorption was comparable to that of DTDSEEEIR-iron, but better than that of DELEDSLER-iron and FeSO4. Thus, peptides with consecutive Glu have the potential as an effective iron carrier to improve iron absorption.

Angiotensin I-Converting Enzyme (ACE-I) Inhibition and Antioxidant Peptide from a Squilla Species.

BACKGROUND: Oratosquilla woodmasoni is one of the marine squilla species, which is found in the entire Asia-Pacific region. This current study assesses the species as the main basis of both ACEi and antioxidant peptide. OBJECTIVE: To isolate the ACEi peptide derived from O. woodmasoni and examine its ACE inhibition along with antioxidant potential. MATERIALS AND METHODS: The squilla muscle protein was hydrolysed using alcalase and trypsin enzymes for 12 hours and tested for DH. The hydrolysates were examined for their ACEi activity and then the best hydrolysate was sequentially purified in various chromatographical methods. The purified peptide was studied for anti-oxidant and functional properties, followed by amino acid sequencing. The purified peptide was also evaluated for its toxicity by in vitro cell viability assay. RESULTS: The DH% was found to be 47.13 ± 0.72% and 89.43 ± 2.06% for alcalase and trypsin, respectively. The alcalase 5th-hour hydrolysate was detected with potent activity (65.97 ± 0.56%) using ACEi assay and was primarily fractionated using ultrafiltration; the maximum inhibitory activity was found with 77.04 ± 0.52% in 3-10 kDa fraction. Subsequently, the fraction was purified using IEC and GFC, in which the AC1-A2 fraction had higher antihypertensive activity (70.85 ± 0.78%). The non-toxic fraction showed hexapeptide HVGGCG with molecular weight 529 Da with great potential of antioxidant activity along with functional property. CONCLUSION: This peptide could be developed as a potential ACE-inhibitory and antioxidant agent.

Biochemical and cytotoxic evaluation of latroeggtoxin-VI against PC12 cells.

Latroeggtoxin-VI (LETX-VI) is a peptide neurotoxin discovered from Latrodectus tredecimguttatus eggs. In the current study, the action features of the neurotoxin on PC12 cells were systematically investigated. LETX-VI could promote dopamine release from PC12 cells in the absence and presence of Ca2+, involving an even more complex action mechanism in the presence of Ca2+ and when the treatment time was longer. Although LETX-VI enchanced the autophagy and secretion activity in PC 12 cells, it showed no remarkable influence on the proliferation, cell cycle, apoptosis and ultrastructure of the cells. Pulldown combined with CapLC-MS/MS analysis suggested that LETX-VI affected PC12 cells by interacting with multiple proteins involved in the metabolism, transport, and release of neurotransmitters, particularly dopamine. The low cytotoxicity and effective regulatory action of LETX-VI on PC12 cells suggest the potential of the active peptide in the development of drugs for the treatment of some dopamine-related psychotic diseases and cancers.

Production of an invertebrate lysozyme of Scylla paramamosain in E.coli and evaluation of its antibacterial, antioxidant and anti-inflammatory effects.

Lysozymes, which are secreted in many organisms, including invertebrates, mammals, plants, bacteria and fungus, exhibit antimicrobial, antiviral, antioxidant, and anti-inflammatory activities. Splys-i is an invertebrate-type (i-type) lysozyme isolated from Scylla paramamosain in 2017 and is involved in immune defense against bacteria. However, the antibacterial, antioxidant, and anti-inflammatory activities of Splys-i remain to be elucidated. In the current study, the expression parameters (including IPTG concentration, induction temperature, and induction duration) of Splys-i in Escherichia coli were optimized to achieve high-level yield through shake-flask cultivation with approximately 120 mg of Splys-i obtained from 1 L of LB medium. The purified Splys-i displayed low cytotoxicity to RAW264.7 macrophage cells and low hemolytic activity against erythrocytes of mouse, rat, and rabbit, respectively, and exhibited potent antibacterial activity against both Gram-positive and -negative bacteria with minimum concentrations ranging from 15 to 90 µg/mL. The antibacterial property of Splys-i was also unaffected when treated with various temperature, pHs, and salinity, respectively, and Splys-i showed resistance to proteinase digestion. Radical-scavenging rate assay (including ABTS+, DPPH, hydroyl free radical, and superoxide anion) indicated that Splys-i was an efficient antioxidant. Splys-i also exerted anti-inflammatory effect through the inhibition of IκBα and NF-κB(P65) phosphorylation, thereby reducing the secretion of pro-inflammatory cytokines. All these results suggested that Splys-i can be prepared from E. coli with potent biological property.

LP2, the first lanthipeptide GPCR agonist in a human pharmacokinetics and safety study.

Introduction of a lanthionine into a peptide may enhance target affinity, target specificity and proteolytic resistance. This manuscript reports preclinical safety studies and the first-in-human study with the lanthipeptide AT2R agonist LP2, a structural analog of cAng-(1-7), whose N-terminus was protected against aminopeptidases by the presence of a d-lysine. None of the preclinical studies, including an in vitro multitarget panel, behavioral, respiratory and cardiovascular measurements, genotoxicity and toxicity studies in rat and dog, posed any safety concern. Due to lack of toxicity the maximum tolerated dose was not reached neither in rat nor in dog. In the human dose escalation study, healthy male volunteers received a single 1 mL subcutaneous injection (0.001 mg, 0.01 mg or 0.1 mg) of LP2 or matching placebo. In contrast to angiotensin II which has a T1/2 in plasma of < 1 min, LP2 has a T1/2 of approximately 2.1-2.6 hours. The fraction of the dose excreted unchanged in urine ranged from 84.73 ± 10.4 % at a dose of 0.001 mg to 66.4 ± 3.9 % at 0.1 mg. There were no deaths, serious adverse events or subject withdrawals as a result of an adverse event. The incidence of adverse events was 16.7 %; each was mild in severity. One adverse event, peripheral coldness, was considered to be possibly related to LP2 at 0.001 mg LP2. None of the results was considered to pose a clinically relevant safety concern. This study supports the potential for the therapeutic use of lanthipeptides.

Structural modelling and thermostability of a serine protease inhibitor belonging to the Kunitz-BPTI family from the Rhipicephalus microplus tick.

rBmTI-A is a recombinant serine protease inhibitor that belongs to the Kunitz-BPTI family and that was cloned from Rhipicephalus microplus tick. rBmTI-A has inhibitory activities on bovine trypsin, human plasma kallikrein, human neutrophil elastase and plasmin with dissociation constants in nM range. It is characterized by two inhibitory domains and each domain presents six cysteines that form three disulfide bonds, which contribute to the high stability of its structure. Previous studies suggest that serine protease inhibitor rBmTI-A has a protective potential against pulmonary emphysema in mice and anti-inflammatory potential. Besides that, rBmTI-A presented a potent inhibitory activity against in vitro vessel formation. In this study, the tertiary structure of rBmTI-A was modeled. The structure stabilization was evaluated by molecular dynamics analysis. Circular dichroism spectroscopy data corroborated the secondary structure found by the homology modelling. Also, in circular dichroism data it was shown a thermostability of rBmTI-A until approximately 70 °C, corroborated by inhibitory assays toward trypsin.

Cyclic gomesin, a stable redesigned spider peptide able to enter cancer cells.

Anticancer chemo- and targeted therapies are limited in some cases due to strong side effects and/or drug resistance. Peptides have received renascent interest as anticancer therapeutics and are currently being considered as alternatives and/or as complementary to biologics and small-molecule drugs. Gomesin, a disulfide-rich host defense peptide expressed in the Brazilian spider Acanthoscurria gomesiana selectively targets and disrupts cancer cell membranes. In the current study, we employed a range of biophysical methodologies with model membranes and bioassays to investigate the use of a cyclic analogue of gomesin as a drug scaffold to internalize cancer cells. We found that cyclic gomesin can internalize cancer cells via endocytosis and direct membrane permeation. In addition, we designed an improved non-disruptive and non-toxic cyclic gomesin analogue by incorporating D-amino acids within the scaffold. This improved analogue retained the ability to enter cancer cells and can be used as a scaffold to deliver drugs. Efforts to investigate the internalization mechanism used by host defense peptides, and to improve their stability, potency, selectivity and ability to permeate cancer cell membranes will increase the opportunities to repurpose peptides as templates for designing alternative anticancer therapeutic leads.

Arg-substituted VmCT1 analogs reveals promising candidate for the development of new antichagasic agent.

VmCT1 is an antimicrobial peptide (AMP) isolated from the venom of the scorpion Vaejovis mexicanus with antimicrobial, anticancer and antimalarial activities, which the rational design with Arg-substitution has yielded AMPs with higher antimicrobial activity than VmCT1. Chagas is a neglected tropical disease, becoming the development of new antichagasic agents is urgent. Thus, we aimed to evaluate the antichagasic effect of VmCT1 and three Arg-substituted analogues, as well their action mechanism. Peptides were tested against the epimastigote, trypomastigote, amastigote forms of Trypanossoma cruzi Y strain and against LLC-MK2 mammalian cells. The mechanism of action of these peptides was evaluated by means of flow cytometry and scanning electron microscopy. VmCT1 presented activity against all three forms of T. cruzi, with EC50 against trypomastigote forms of 1.37 µmol L-1 and selectivity index (SI) of 58. [Arg]3-VmCT1, [Arg]7-VmCT1 and [Arg]11-VmCT1 also showed trypanocidal effect, but [Arg]11-VmCT1 had the best effect, being able to decrease the EC50 against trypomastigote forms to 0.8 µmol L-1 and increase SI to 175. Necrosis was cell death pathway of VmCT1, as well [Arg]7-VmCT1 and [Arg]11-VmCT1, such as observed by membrane damage in flow cytometry analyses and scanning-electron-microscopy. In conclusion, [Arg]11-VmCT1 revealed promising as a candidate for new antichagasic therapeutics.

Biological activities of a recombinant fortilin from Fenneropenaeus merguiensis.

Human Fortilin, an antiapoptotic protein, has also been implicated in several diseases; however, several potential uses of fortilin have also been proposed. Bearing the implications of fortilin in mind, fortilin analog, which has no complication with diseases, is required. Since a recombinant full-length fortilin from Fenneropenaeus merguiensis (rFm-Fortilin (FL)) reported only 44% (3e-27) homologous to human fortilin, therefore the biological activities of the Fm-Fortilin (FL) and its fragments (F2, F12, and F23) were investigated for potential use against HEMA toxicity from filling cement to pulp cell. The rFm-Fortilin FL, F2, 12, and F23 were expressed and assayed for proliferation activity. The rFm-Fortilin (FL) showed proliferation activity on human dental pulp cells (HDPCs) and protected the cells from 2-hydroxy-ethyl methacrylate (HEMA) at 1-20 ng/ml. In contrast, none of the rFm-Fortilin fragments promoted HDPC growth that may be due to a lack of three conserved amino acid residues together for binding with the surface of Rab GTPase for proliferative activity. In addition, rFm-Fortilin (FL) activated mineralization and trend to suppressed production of proinflammatory cytokines, including histamine (at 10 ng/ml) and TNF-α (at 100 ng/ml). Besides, the rFm-Fortilin (FL) did not mutate the Chinese hamster ovary (CHO) cell. Therefore, the rFm-Fortilin (FL) has the potential use as a supplementary medical material to promote cell proliferation in patients suffering severe tooth decay and other conditions.

Proteotranscriptomic Insights into the Venom Composition of the Wolf Spider Lycosa tarantula.

Spider venoms represent an original source of novel compounds with therapeutic and agrochemical potential. Whereas most of the research efforts have focused on large mygalomorph spiders, araneomorph spiders are equally promising but require more sensitive and sophisticated approaches given their limited size and reduced venom yield. Belonging to the latter group, the genus Lycosa ("wolf spiders") contains many species widely distributed throughout the world. These spiders are ambush predators that do not build webs but instead rely strongly on their venom for prey capture. Lycosa tarantula is one of the largest species of wolf spider, but its venom composition is unknown. Using a combination of RNA sequencing of the venom glands and venom proteomics, we provide the first overview of the peptides and proteins produced by this iconic Mediterranean spider. Beside the typical small disulfide rich neurotoxins, several families of proteins were also identified, including cysteine-rich secretory proteins (CRISP) and Hyaluronidases. Proteomic analysis of the electrically stimulated venom validated 30 of these transcriptomic sequences, including nine putative neurotoxins and eight venom proteins. Interestingly, LC-MS venom profiles of manual versus electric stimulation, as well as female versus male, showed some marked differences in mass distribution. Finally, we also present some preliminary data on the biological activity of L. tarantula crude venom.

Scorpion Toxins and Ion Channels: Potential Applications in Cancer Therapy.

Apoptosis, a genetically directed process of cell death, has been studied for many years, and the biochemical mechanisms that surround it are well known and described. There are at least three pathways by which apoptosis occurs, and each pathway depends on extra or intracellular processes for activation. Apoptosis is a vital process, but disturbances in proliferation and cell death rates can lead to the development of diseases like cancer. Several compounds, isolated from scorpion venoms, exhibit inhibitory effects on different cancer cells. Indeed, some of these compounds can differentiate between healthy and cancer cells within the same tissue. During the carcinogenic process, morphological, biochemical, and biological changes occur that enable these compounds to modulate cancer but not healthy cells. This review highlights cancer cell features that enable modulation by scorpion neurotoxins. The properties of the isolated scorpion neurotoxins in cancer cells and the potential uses of these compounds as alternative treatments for cancer are discussed.

Amblyomin-X, a recombinant Kunitz-type inhibitor, regulates cell adhesion and migration of human tumor cells.

In a tumor microenvironment, endothelial cell migration and angiogenesis allow cancer to spread to other organs causing metastasis.  Indeed, a number of molecules that are involved in cytoskeleton re-organization and intracellular signaling have been investigated for their effects on tumor cell growth and metastasis. Alongside that, Amblyomin-X, a recombinant Kunitz-type protein, has been shown to reduce metastasis and tumor growth in in vivo experiments. In the present report, we provide a mechanistic insight to these antitumor effects, this is,  Amblyomin-X modulates Rho-GTPases and uPAR signaling, and reduces the release of MMPs, leading to disruption of the actin cytoskeleton and decreased cell migration of tumor cell lines. Altogether, our data support a role for Amblyomin-X as a novel potential antitumor drug. ABBREVIATIONS: Amb-X: Amblyomin-X; ECGF: endotelial cell growth factor; ECM: extracellular matrix; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HUVEC: human umbilical vein endothelial cell; LRP1: low-density lipoprotein receptor-related protein; MMP: matrix metalloproteinase; HPI-4: hedgehog pathway inhibitor 4; PAI-1: plasminogen activator inhibitor 1; PMA: phorbol 12-myristate-13-acetate; TFPI: tissue factor pathway inhibitor; uPA: urokinase plasminogen activator; uPAR: uPA receptor.

The immunosuppressive functions of two novel tick serpins, HlSerpin-a and HlSerpin-b, from Haemaphysalis longicornis.

Serpins are evolutionarily conserved serine protease inhibitors that are widely distributed in animals, plants and microbes. In this study, we reported the cloning and functional characterizations of two novel serpin genes, HlSerpin-a and HlSerpin-b, from the hard tick Haemaphysalis longicornis of China. Recombinant HlSerpin-a and HlSerpin-b displayed protease inhibitory activities against multiple mammalian proteases. Similar to other tick serpins, HlSerpin-a and HlSerpin-b suppressed the expression of inflammatory cytokines such as TNF-α, interleukin (IL)-6 and IL-1ß from lipopolysaccharide-stimulated mouse bone-marrow-derived macrophages (BMDMs) or mouse bone-marrow-derived dendritic cells (BMDCs). The minimum active region (reaction centre loop) of HlSerpin-a, named SA-RCL, showed similar biological activities as HlSerpin-a in the protease inhibition and immune suppression assays. The immunosuppressive activities of full-length HlSerpin-a and SA-RCL are impaired in Cathepsin G or Cathepsin B knockout mouse macrophages, suggesting that the immunomodulation functions of SA and SA-RCL are dependent on their protease inhibitory activity. Finally, we showed that both full-length HlSerpins and SA-RCL can relieve the joint swelling and inflammatory response in collagen-induced mouse arthritis models. These results suggested that HlSerpin-a and HlSerpin-b are two functional arthropod serpins, and the minimal reactive peptide SA-RCL is a potential candidate for drug development against inflammatory diseases.

Validation of Strasbourg environmental exposure chamber (EEC) ALYATEC® in mite allergic subjects with asthma.

Objective: Environmental Exposure Chamber (EEC) should have standardized and controlled allergenic and non-allergenic exposures to perform reproducible clinical studies. The aim was to demonstrate that mite exposure in the Alyatec® EEC could induce early (EAR) and/or late asthmatic reactions (LAR) in at least 60% of subjects allergic to mite.Methods: The EEC has a volume of 147-m3 with 20 seats. The nebulized particle number, airborne Der p1, endotoxins, and volatile organic compound (VOC) concentrations were measured. Twenty-four asthmatics allergic to mite were randomly exposed to 15, 25, and 46 ng/m3 Der p1. Specificity was assessed in not mite-sensitized asthmatics.Results: No significant endotoxin or VOC contamination was measured. The mean inter-assay CVs were 12.5% for the airborne particle number and 28.7% for airborne Der p1 concentrations. For the three Der p1 concentrations, at least 88% of the subjects developed EAR and/or LAR, and at least 46% developed a dual response. No reaction occurred with placebo or in the control group. No severe bronchial reaction occurred.Conclusions: The Alyatec® EEC demonstrated a tight control of allergenic and non-allergenic exposures. The EEC was clinically validated, with airborne Der p1 levels close to levels found in natural settings.

Biodegradable cross-linked chitosan nanoparticles improve anti-Candida and anti-biofilm activity of TistH, a peptide identified in the venom gland of the Tityus stigmurus scorpion.

Among several bioactive peptides identified from the venom glands of the Tityus stigmurus scorpion, one peptide with hypotensive action (TistH, Tityus stigmurus Hypotensin) showed multifunctional and biotechnological applications. The maximum efficacy of this class of compounds can be achieved by immobilizing it in specific and suitable biomaterials or suitable carriers. In this study, distinct entrapment methods of TistH in chitosan nanoparticles was tested using its incorporation (CN-TistH-Inc) or adsorption (CN-TistH-Ads) methods by ionotropic gelification. Physico-chemical properties as well as biocompatibility and antifungal efficacy were assessed for different samples. Atomic force microscopy and field emission gun scanning electronic microscopy images associated with particle size measurements demonstrated that the two methods induced cationic spherical, small (< 160 nm), and narrow-sized (PdI about 0.3) nanoparticles, even after peptide loading greater than 96.5%, which was confirmed using Fourier transform infrared spectroscopy. The colloidal suspensions showed to be stable for 8 weeks and were able to induce the desired slow in vitro peptide release. Cytotoxicity assays performed in normal cells originated from murine macrophages (RAW 264.7) and kidneys of African green monkeys (Vero E6) suggested biocompatibility of samples. The CN-TistH-Inc and CN-TistH-Ads showed a minimal inhibitory concentration of 89.2 µg.mL-1 against Candida albicans, 11.1 µg.mL-1 for C. parapsilosis and C. tropicalis, confirmed by minimum fungicidal concentrations assay. Moreover, the TistH-loaded cross-linked chitosan nanoparticles significantly reduced the biofilm formation of clinical yeast sepsis of C. tropicalis and C. krusei, as well as clinical yeasts of vulvovaginal candidiasis of C. albicans. In this approach, biodegradable nanocarriers prepared using simple and reproducible methods demonstrated the ability to deliver the TistH peptide from T. stigmurus and improve its antifungal efficacy.

Pore-forming spider venom peptides show cytotoxicity to hyperpolarized cancer cells expressing K+ channels: A lentiviral vector approach.

Recent studies demonstrated the upregulation of K+ channels in cancer cells. We have previously found that a pore-forming peptide LaFr26, purified from the venom of the Lachesana sp spider, was selectively incorporated into K+ channel expressing hyperpolarized cells. Therefore, it is expected that this peptide would have selective cytotoxicity to hyperpolarized cancer cells. Here we have tested whether LaFr26 and its related peptide, oxyopinin-2b, are selectively cytotoxic to K+ channel expressing cancer cells. These peptides were cytotoxic to the cells, of which resting membrane potential was hyperpolarized. The vulnerabilities of K+ channel-expressing cell lines correlated with their resting membrane potential. They were cytotoxic to lung cancer cell lines LX22 and BEN, which endogenously expressed K+ current. Contrastingly, these peptides were ineffective to glioblastoma cell lines, U87 and T98G, of which membrane potentials were depolarized. Peptides have a drawback, i.e. poor drug-delivery, that hinders their potential use as medicine. To overcome this drawback, we prepared lentiviral vectors that can express these pore-forming peptides and tested the cytotoxicity to K+ channel expressing cells. The transduction with these lentiviral vectors showed autotoxic activity to the channel expressing cells. Our study provides the basis for a new oncolytic viral therapy.

Tying pest insects in knots: the deployment of spider-venom-derived knottins as bioinsecticides.

Spider venoms are complex chemical arsenals that contain a rich variety of insecticidal toxins. However, the major toxin class in many spider venoms is disulfide-rich peptides known as knottins. The knotted three-dimensional fold of these mini-proteins provides them with exceptional chemical and thermal stability as well as resistance to proteases. In contrast with other bioinsecticides, which are often slow-acting, spider knottins are fast-acting neurotoxins. In addition to being potently insecticidal, some knottins have exceptional taxonomic selectivity, being lethal to key agricultural pests but innocuous to vertebrates and beneficial insects such as bees. The intrinsic oral activity of these peptides, combined with the ability of aerosolized knottins to penetrate insect spiracles, has enabled them to be developed commercially as eco-friendly bioinsecticides. Moreover, it has been demonstrated that spider-knottin transgenes can be used to engineer faster-acting entomopathogens and insect-resistant crops. © 2019 Society of Chemical Industry.