The Role of Snake Venom Disintegrins in Angiogenesis.

Angiogenesis, the formation of new blood vessels, plays a critical role in various physiological and pathological conditions. Snake venom disintegrins (SVDs) have been identified as significant regulators of this process. In this review, we explore the dual roles of SVD in angiogenesis, both as antiangiogenic agents by inhibiting integrin binding and interfering with vascular endothelial growth factors and as proangiogenic agents by enhancing integrin binding, stimulating cell migration and proliferation, and inducing neoangiogenesis. Studies in vitro and in animal models have demonstrated these effects and offer significant therapeutic opportunities. The potential applications of SVD in diseases related to angiogenesis, such as cancer, ocular diseases, tissue regeneration, wound healing, and cardiovascular diseases, are also discussed. Overall, SVDs are promising potential therapeutics, and further advances in this field could lead to innovative treatments for diseases related to angiogenesis.

The role of snake venom disintegrins in Angiogenesis

Angiogenesis, the formation of new blood vessels, plays a critical role in various physiological and pathological conditions. Snake venom disintegrins (SVDs) have been identified as significant regulators of this process. In this review, we explore the dual roles of SVD in angiogenesis, both as antiangiogenic agents by inhibiting integrin binding and interfering with vascular endothelial growth factors and as proangiogenic agents by enhancing integrin binding, stimulating cell migration and proliferation, and inducing neoangiogenesis. Studies in vitro and in animal models have demonstrated these effects and offer significant therapeutic opportunities. The potential applications of SVD in diseases related to angiogenesis, such as cancer, ocular diseases, tissue regeneration, wound healing, and cardiovascular diseases, are also discussed. Overall, SVDs are promising potential therapeutics, and further advances in this field could lead to innovative treatments for diseases related to angiogenesis.

Snake venom disintegrins update: insights about new findings.

Snake venom disintegrins are low molecular weight, non-enzymatic proteins rich in cysteine, present in the venom of snakes from the families Viperidae, Crotalidae, Atractaspididae, Elapidae, and Colubridae. This family of proteins originated in venom through the proteolytic processing of metalloproteinases (SVMPs), which, in turn, evolved from a gene encoding an A Disintegrin And Metalloprotease (ADAM) molecule. Disintegrins have a recognition motif for integrins in their structure, allowing interaction with these transmembrane adhesion receptors and preventing their binding to proteins in the extracellular matrix and other cells. This interaction gives disintegrins their wide range of biological functions, including inhibition of platelet aggregation and antitumor activity. As a result, many studies have been conducted in an attempt to use these natural compounds as a basis for developing therapies for the treatment of various diseases. Furthermore, the FDA has approved Tirofiban and Eptifibatide as antiplatelet compounds, and they are synthesized from the structure of echistatin and barbourin, respectively. In this review, we discuss some of the main functional and structural characteristics of this class of proteins and their potential for therapeutic use.

ADAMTS-7 Modulates Atherosclerotic Plaque Formation by Degradation of TIMP-1.

BACKGROUND: The ADAMTS7 locus was genome-wide significantly associated with coronary artery disease. Lack of the ECM (extracellular matrix) protease ADAMTS-7 (A disintegrin and metalloproteinase-7) was shown to reduce atherosclerotic plaque formation. Here, we sought to identify molecular mechanisms and downstream targets of ADAMTS-7 mediating the risk of atherosclerosis. METHODS: Targets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, coimmunoprecipitation, and Förster resonance energy transfer-based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. RESULTS: In humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe-/- mice, atherosclerotic aortas of Apoe-/- mice lacking Adamts-7 (Apoe-/-Adamts7-/-) contained higher protein levels of Timp-1 (tissue inhibitor of metalloprotease-1). In coimmunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target MMP-9 (matrix metalloprotease-9). As a downstream mechanism, we investigated collagen content in plaques of Apoe-/- and Apoe-/-Adamts7-/- mice after a Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. To facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim of decreasing TIMP-1 degradation, we designed a Förster resonance energy transfer-based assay targeting the ADAMTS-7 catalytic site. CONCLUSIONS: ADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise associated with coronary artery disease. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for the reduction of atherosclerosis-related events.

γ-Bungarotoxin impairs the vascular endothelial barrier function by inhibiting integrin α5.

γ-bungarotoxin (γ-BGT) is an RGD motif-containing protein, derived from the venom of Bungarus multicinctus, leading to acute death in mice. These RGD motif-containing proteins from snake venom belonging to the disintegrin family can interfere with vascular endothelial homeostasis by directly binding cell surface integrins. Targeting integrins that generate vascular endothelial dysfunction may contribute to γ-BGT poisoning, however, the underlying mechanisms have not been investigated in detail. In this study, the results showed that γ-BGT played a role in -promoting the permeability of the vascular endothelial barrier. Depending on its selective binding to integrin α5 in vascular endothelium (VE), γ-BGT initiated downstream events, including focal adhesion kinase dephosphorylation and cytoskeleton remodeling, resulting in the intercellular junction interruption. Those alternations facilitated paracellular permeability of VE and barrier dysfunction. Proteomics profiling identified that as a downstream effector of the integrin α5 / FAK signaling pathway cyclin D1 partially mediated the cellular structural changes and barrier dysfunction. Furthermore, VE-released plasminogen activator urokinase and platelet-derived growth factor D could serve as potential diagnostic biomarkers for γ-BGT-induced vascular endothelial dysfunction. Our results indicate the mechanisms through which γ-BGT as a novel disintegrin directly interacts with the VE, with consequences for barrier dysfunction.

Full sequencing and comparison of five venom metalloproteases of Trimeresurus gracilis: The PI-enzyme is most similar to okinalysin but the PIII-enzyme is most similar to Crotalus venom enzymes.

The cDNAs encoding the Zn+2-metalloproteases (SVMPs) of Trimeresurus gracilis (abbreviated as Tgc), a pitviper endemic to Taiwan, were cloned from venom glands and sequenced. The amino-acid sequences of five novel SVMPs, including one P-III, three P-II and one P-I class enzymes, were thus deduced and subjected to BLAST-analyses. The P-III enzyme (designated as Tgc-PIII) is structurally most similar to the PIII-SVMPs of New World pitvipers but not similar to the PIII-SVMP of Ovophis okinavensis. Sequence-similarity analysis of 22 homologous PIII-SVMPs reveal three major structural subtypes of the pitviper PIII-SVMPs, which possibly have different substrate specificities. In addition, Tgc-PIII and the PI-class SVMP (named Tgc-MP) were isolated from the venom and verified by mass spectrometry. All the three deduced sequences of PII-SVMPs (Tgc-PIIs) contain an abnormal Zn+2-binding-site in their catalytic-domain, and an identical "long-disintegrin" domain. The predicted 85-residues disintegrin, gracilisin, bears high similarities to some long-disintegrins of the New-World pitvipers and salmosin3. By BLAST search and comparison, Tgc-MP is 96% similar to okinalysin, the hemorrhagic PI-SVMP of O. okinavensis, rather than any other PI-SVMPs in the databanks. Our results confirm the fast evolution of Tgc-SVMPs as well as their structural similarities to different SVMP-classes of the New-World pitvipers and of O. okinavensis, respectively. The implications of our findings are discussed along with our previous sequence comparisons of venom phospholipases A2 and ten venom serine proteases of Tgc.

How snake venom disintegrins affect platelet aggregation and cancer proliferation.

Disintegrins are small peptides possessing a tripeptide motif capable of binding to integrins. These were first isolated from viper venoms and are now also found in many other hematophagous organisms. Many integrins have been studied for their role in the onset of disease and the interaction of disintegrins with these receptors makes them potential therapeutic molecules. Disintegrins are also used as molecular scaffolds to design effective drugs for cardiovascular diseases and cancer. Even the gene and protein sequencing data of disintegrins have provided insights into understanding the molecular complexity of disintegrins. In this review, we try to summarize the structural and functional importance of disintegrins in identifying the biological targets and triggering various signaling pathways involved in platelet aggregation and cancer. Also, we have tried to elucidate a possible molecular mechanism behind the action of disintegrins on platelet aggregation and cancer. This understanding will help us to design and to explore more of these integrin-binding molecules.

Snake venom disintegrins update: insights about new findings

ABSTRACT Snake venom disintegrins are low molecular weight, non-enzymatic proteins rich in cysteine, present in the venom of snakes from the families Viperidae, Crotalidae, Atractaspididae, Elapidae, and Colubridae. This family of proteins originated in venom through the proteolytic processing of metalloproteinases (SVMPs), which, in turn, evolved from a gene encoding an A Disintegrin And Metalloprotease (ADAM) molecule. Disintegrins have a recognition motif for integrins in their structure, allowing interaction with these transmembrane adhesion receptors and preventing their binding to proteins in the extracellular matrix and other cells. This interaction gives disintegrins their wide range of biological functions, including inhibition of platelet aggregation and antitumor activity. As a result, many studies have been conducted in an attempt to use these natural compounds as a basis for developing therapies for the treatment of various diseases. Furthermore, the FDA has approved Tirofiban and Eptifibatide as antiplatelet compounds, and they are synthesized from the structure of echistatin and barbourin, respectively. In this review, we discuss some of the main functional and structural characteristics of this class of proteins and their potential for therapeutic use.

15N, 13C, and 1H resonance assignments of Jarastatin: a disintegrin of Bothrops jararaca.

Disintegrins are a group of cysteine-rich proteins found in a wide variety of snake venoms. These proteins selectively bind to integrins, which play a fundamental role in the regulation of many physiological and pathological processes. Here, we report the NMR chemical shift assignments for 1H, 15N, and 13C nuclei in the backbone and side chains of recombinant disintegrin Jarastatin (rJast), which was further validated by secondary structure prediction using the TALOS-N server. Taken together, these data are essential to perform NMR-based experiments, including structure determination, backbone dynamics, mapping ligand sites and enabling a deeper understanding of the effect of hydrophobic surface clusters, which are important elements to stabilize some 3D proteins structure/folding.

The First Snake Venom KTS/Disintegrins-Integrin Interactions Using Bioinformatics Approaches.

Snake venom contains a number of active molecules that have been shown to possess high anti-tumor activities; disintegrins are an excellent example among these. Their ability to interact and bind with integrins suggests that they could be very valuable molecules for the development of new cancer therapeutic approaches. However, in the absence of a clear Lysine-Threonine-Serine (KTS) Disintegrins Integrin interaction model, the exact compound features behind it are still unknown. In this study, we investigated the structural characteristics of three KTS-disintegrins and the interaction mechanisms with the α1ß1 integrin receptor using in silico bioinformatics approaches. Normal mode analysis showed that the flexibility of the KTSR motif and the C-terminal region play a key role and influence the KTS-Disintegrin-integrin interaction. Protein-protein docking also suggested that the interaction involving the KTSR motif is highly dependent on the residue following K21, S23 and R24. These findings contribute to a better understanding of the KTS-Disintegrin-Integrin structural differences and their interactions with α1ß1 receptors, which could improve the selection process of the best active molecules for antitumor therapies.

Immunorecognition capacity of Indian polyvalent antivenom against venom toxins from two populations of Echis carinatus.

Clinicians report low efficacy of Indian polyvalent antivenom (PAV), with >20 vials required for treatment of a snakebite envenoming. We hypothesize that the antivenom efficacy could be reduced due to insufficient antibodies against some venom toxins. To test this, we used third-generation antivenomics to reveal bound and unbound venom toxins of Echis carinatus venom from Goa (ECVGO) and Tamil Nadu (ECVTN). We used 60, 120, 180, 240, 300, and 360 µg of venom and passed through mini-columns containing ~5 mg Antivenom bound to CNBr beads. The non-retained (unbound) and retained (bound) toxins were identified using reverse-phase HPLC and tandem mass spectrometry. Low molecular weight toxins - Short disintegrins (5.3 kDa) and DIS domain of P-II SVMP from ECVGO and ECVTN showed poor binding with antivenom. The immunorecognition sites of antivenom saturated at the lower antivenom-venom ratio for ECVGO than for ECVTN. The immunoretained capacity of antivenom against ECVTN was 140.6 µg and ECVGO was 125.1 µg. The amount of immunoretained toxins quantified can further be used to estimate the efficacy of antivenom by correlating it with in-vivo studies. The unbound toxins identified from this study could be targeted to improve the effectiveness of antivenom.

Effects of venoms on neutrophil respiratory burst: a major inflammatory function.

Neutrophils play a pivotal role in innate immunity and in the inflammatory response. Neutrophils are very motile cells that are rapidly recruited to the inflammatory site as the body first line of defense. Their bactericidal activity is due to the release into the phagocytic vacuole, called phagosome, of several toxic molecules directed against microbes. Neutrophil stimulation induces release of this arsenal into the phagosome and induces the assembly at the membrane of subunits of the NAPDH oxidase, the enzyme responsible for the production of superoxide anion that gives rise to other reactive oxygen species (ROS), a process called respiratory burst. Altogether, they are responsible for the bactericidal activity of the neutrophils. Excessive activation of neutrophils can lead to extensive release of these toxic agents, inducing tissue injury and the inflammatory reaction. Envenomation, caused by different animal species (bees, wasps, scorpions, snakes etc.), is well known to induce a local and acute inflammatory reaction, characterized by recruitment and activation of leukocytes and the release of several inflammatory mediators, including prostaglandins and cytokines. Venoms contain several molecules such as enzymes (phospholipase A2, L-amino acid oxidase and proteases, among others) and peptides (disintegrins, mastoporan, parabutoporin etc.). These molecules are able to stimulate or inhibit ROS production by neutrophils. The present review article gives a general overview of the main neutrophil functions focusing on ROS production and summarizes how venoms and venom molecules can affect this function.

Effects of venoms on neutrophil respiratory burst: a major inflammatory function

Neutrophils play a pivotal role in innate immunity and in the inflammatory response. Neutrophils are very motile cells that are rapidly recruited to the inflammatory site as the body first line of defense. Their bactericidal activity is due to the release into the phagocytic vacuole, called phagosome, of several toxic molecules directed against microbes. Neutrophil stimulation induces release of this arsenal into the phagosome and induces the assembly at the membrane of subunits of the NAPDH oxidase, the enzyme responsible for the production of superoxide anion that gives rise to other reactive oxygen species (ROS), a process called respiratory burst. Altogether, they are responsible for the bactericidal activity of the neutrophils. Excessive activation of neutrophils can lead to extensive release of these toxic agents, inducing tissue injury and the inflammatory reaction. Envenomation, caused by different animal species (bees, wasps, scorpions, snakes etc.), is well known to induce a local and acute inflammatory reaction, characterized by recruitment and activation of leukocytes and the release of several inflammatory mediators, including prostaglandins and cytokines. Venoms contain several molecules such as enzymes (phospholipase A2, L-amino acid oxidase and proteases, among others) and peptides (disintegrins, mastoporan, parabutoporin etc.). These molecules are able to stimulate or inhibit ROS production by neutrophils. The present review article gives a general overview of the main neutrophil functions focusing on ROS production and summarizes how venoms and venom molecules can affect this function.(AU)

Structural Insight into Integrin Recognition and Anticancer Activity of Echistatin.

Echistatin (Ech) is a short disintegrin with a long 42NPHKGPAT C-terminal tail. We determined the 3-D structure of Ech by X-ray crystallography. Superimposition of the structures of chains A and B showed conformational differences in their RGD loops and C-termini. The chain A structure is consistent with our NMR analysis that the GPAT residues of the C-terminus cannot be observed due to high flexibility. The hydrogen bond patterns of the RGD loop and between the RGD loop and C-terminus in Ech were the same as those of the corresponding residues in medium disintegrins. The mutant with C-terminal HKGPAT truncation caused 6.4-, 7.0-, 11.7-, and 18.6-fold decreases in inhibiting integrins αvß3, αIIbß3, αvß5, and α5ß1. Mutagenesis of the C-terminus showed that the H44A mutant caused 2.5- and 4.4-fold increases in inhibiting αIIbß3 and α5ß1, and the K45A mutant caused a 2.6-fold decrease in inhibiting αIIbß3. We found that Ech inhibited VEGF-induced HUVEC proliferation with an IC50 value of 103.2 nM and inhibited the migration of A375, U373MG, and Panc-1 tumor cells with IC50 values of 1.5, 5.7, and 154.5 nM. These findings suggest that Ech is a potential anticancer agent, and its C-terminal region can be optimized to improve its anticancer activity.

Preclinical studies of a novel snake venom-derived recombinant disintegrin with antitumor activity: A review.

Snake venoms consist of a complex mixture of many bioactive molecules. Among them are disintegrins, which are peptides without enzymatic activity, but with high binding affinity for integrins, transmembrane receptors that function to connect cells with components of the extracellular matrix. Integrin-mediated cell attachment is critical for cell migration and dissemination, as well as for signal transduction pathways involved in cell growth. During tumor development, integrins play key roles by supporting cancer cell proliferation, angiogenesis, and metastasis. The recognition that snake venom disintegrins can block integrin functions has spawned a number of studies to explore their cancer therapeutic potential. While dozens of different disintegrins have been isolated, none of them as yet has undergone clinical evaluation in cancer patients. Among the best-characterized and preclinically most advanced disintegrins is vicrostatin (VCN), a recombinant disintegrin that was rationally designed by fusing 62 N-terminal amino acids derived from the disintegrin contortrostatin with 6 C-terminal amino acids from echistatin, the disintegrins from another snake species. Bacterially produced VCN was shown to target multiple tumor-associated integrins, achieving potent anti-tumor and anti-angiogenic effects in in vitro and in vivo models in the absence of noticeable toxicity. This review will introduce the field of snake venom disintegrins as potential anticancer agents and illustrate the translational development and cancer-therapeutic potential of VCN as an example.

Improved Antithrombotic Activity and Diminished Bleeding Side Effect of a PEGylated αIIbß3 Antagonist, Disintegrin.

Polymer polyethylene glycol (PEG), or PEGylation of polypeptides improves protein drug stability by decrease degradation and reduces renal clearance. To produce a pharmaceutical disintegrin derivative, the N-terminal PEGylation technique was used to modify the disintegrin derivative [KGDRR]trimucrin for favorable safety and pharmacokinetic profiles and antithrombotic efficacy. We compared intact [KGDRR]trimucrin (RR) and PEGylated KGDRR (PEG-RR) by in vitro and in vivo systems for their antithrombotic activities. The activity of platelet aggregation inhibition and the bleeding tendency side effect were also investigated. PEG-RR exhibited optimal potency in inhibiting platelet aggregation of human/mouse platelet-rich plasma activated by collagen or ADP with a lower IC50 than the intact derivative RR. In the illumination-induced mesenteric venous thrombosis model, RR and PEG-RR efficaciously prevented occlusive thrombosis in a dose-dependent manner. In rotational thromboelastometry assay, there was no effect of PEG-RR in human whole blood coagulation even given at a higher concentration (30 µg/mL), while RR slightly prolonged clotting time. However, RR and PEG-RR were not associated with severe thrombocytopenia or bleeding in FcγRIIa-transgenic mice at equally efficacious antithrombotic dosages. We also found the in vivo half-life of PEGylation was longer than RR (RR: 15.65 h vs. PEG-RR: 20.45 h). In conclusion, injectable PEG-RR with prolonged half-life and decreased bleeding risk is a safer anti-thrombotic agent for long-acting treatment of thrombus diseases.

ADAM (a Disintegrin and Metalloproteinase) 15 Deficiency Exacerbates Ang II (Angiotensin II)-Induced Aortic Remodeling Leading to Abdominal Aortic Aneurysm.

OBJECTIVE: ADAM (a disintegrin and metalloproteinase) 15-a membrane-bound metalloprotease from the ADAM (disintegrin and metalloproteinase) family-has been linked to endothelial permeability, inflammation, and metastasis. However, its function in aortic aneurysm has not been explored. We aimed to determine the function of ADAM15 in the pathogenesis of aortic remodeling and aneurysm formation. Approach and Results: Male Adam15-deficient and WT (wild type) mice (10 weeks old), on standard laboratory diet, received Ang II (angiotensin II; 1.5 mg/kg per day) or saline (Alzet pump) for 2 or 4 weeks. Ang II increased ADAM15 in WT aorta, while Adam15-deficiency resulted in abdominal aortic aneurysm characterized by loss of medial smooth muscle cells (SMCs), elastin fragmentation, inflammation, but unaltered Ang II-mediated hypertension. In the abdominal aortic tissue and primary aortic SMCs culture, Adam15 deficiency decreased SMC proliferation, increased apoptosis, and reduced contractile properties along with F-actin depolymerization to G-actin. Ang II triggered a markedly greater increase in THBS (thrombospondin) 1 in Adam15-deficient aorta, primarily the medial layer in vivo, and in aortic SMC in vitro; increased SSH1 (slingshot homolog 1) phosphatase activity and cofilin dephosphorylation that promoted F-actin depolymerization and G-actin accumulation. rhTHBS1 (recombinant THBS1) alone was sufficient to activate the cofilin pathway, increase G-actin, and induce apoptosis of aortic SMCs, confirming the key role of THBS1 in this process. Further, in human abdominal aortic aneurysm specimens, decreased ADAM15 was associated with increased THBS1 levels and loss of medial SMCs. CONCLUSIONS: This study is the first to demonstrate a key role for ADAM15 in abdominal aortic aneurysm through regulating the SMC function, thereby placing ADAM15 in a critical position as a potential therapeutic target for abdominal aortic aneurysm.

Syzygium Polyanthum Reduced TNF-α and ADAM17 Protein Expression in Myocardial Infarction Rat Model.

BACKGROUND: In myocardial infarction (MI), inflammatory cytokine including tumor necrosis factor-α (TNF-α) plays pivotal role throughout worsening processes and recovery, whereas its cleavage is controlled by a disintegrin and metalloproteinases-17 (ADAM-17). Syzygium polyanthum (Wight) is widely used plant in Indonesia and Southeast Asian countries to treat various illnesses. OBJECTIVE: This study aimed to analyze the effect of S. polyanthum extract towards TNF-α and ADAM17 expressions in MI rat model. METHODS: Myocardial infarction were surgically induced in wistar rats by ligating left anterior descending coronary artery in both treatment and control group. Around 3.6 mg of S. polyanthum extract (SPE) was administered via nasogastric tube to treatment groups, while control group received only saline. Serum TNF-α level and expression of TNF-α and ADAM17 in blanching heart muscle was examined in both groups on day 1, day 4, day 7, and day 14 after treatment with SPE. RESULTS: Reduction of serum TNF-α levels was markedly seen from day 4 in treatment group and was lower than in control group since day 4. Expression of ADAM17 was reduced and expression of TNF-α was only seen in myocardial membrane in group with SPE treatment. CONCLUSION: Syzygium polyanthum exerts its anti-inflammatory effect by decreasing ADAM17 expression subsequently lowering TNF-α regulation in MI rat model.

Methods for Evaluation of a Snake Venom-Derived Disintegrin in Animal Models of Human Cancer.

Integrin targeting has been shown to be an effective approach for anticancer therapy. We engineered a recombinant disintegrin, vicrostatin (VCN), that binds with high affinity and specificity to the Arg-Gly-Asp (RGD) class of integrins, including αvß3, αvß5, and α5ß1, involved in tumor invasion and metastasis. We used three different delivery modalities to examine anticancer activity of VCN in mouse models of human ovarian cancer, glioma, and prostate cancer. A female mouse model was used to examine the treatment of established ovarian cancer (OC) using VCN delivered intraperitoneally (IP) weekly either in saline or impregnated in a viscoelastic gel. SKOV3luc cells (a human OC cell line) were directly injected IP into immunodeficient mice. We also examined the antitumor activity of radioiodinated VCN delivered intravenously in a human glioma model in nude mice. We evaluated the effectiveness of 131I-VCN in combination with the DNA alkylating agent temozolomide in limiting glioma growth. Finally, treatment of a bone metastatic model of human prostate cancer (PC) in immunodeficient mice was examined using a liposomal formulation of VCN (LVCN) delivered intravenously. Human PC cells were suspended in a solution of Matrigel and injected into the left tibia of immunodeficient mice. Diameters of both the left and right (control) tibias were measured by caliper repeatedly after VCN treatment was initiated.

A Novel αIIbß3 Antagonist from Snake Venom Prevents Thrombosis without Causing Bleeding.

Life-threatening thrombocytopenia and bleeding, common side effects of clinically available αIIbß3 antagonists, are associated with the induction of ligand-induced integrin conformational changes and exposure of ligand-induced binding sites (LIBSs). To address this issue, we examined intrinsic mechanisms and structure-activity relationships of purified disintegrins, from Protobothrops flavoviridis venom (i.e., Trimeresurus flavoviridis), TFV-1 and TFV-3 with distinctly different pro-hemorrhagic tendencies. TFV-1 with a different αIIbß3 binding epitope from that of TFV-3 and chimeric 7 × 103 Fab, i.e., Abciximab, decelerates αIIbß3 ligation without causing a conformational change in αIIbß3, as determined with the LIBS antibody, AP5, and the mimetic, drug-dependent antibody (DDAb), AP2, an inhibitory monoclonal antibody raised against αIIbß3. Consistent with their different binding epitopes, a combination of TFV-1 and AP2 did not induce FcγRIIa-mediated activation of the ITAM-Syk-PLCγ2 pathway and platelet aggregation, in contrast to the clinical antithrombotics, abciximab, eptifibatide, and disintegrin TFV-3. Furthermore, TFV-1 selectively inhibits Gα13-mediated platelet aggregation without affecting talin-driven clot firmness, which is responsible for physiological hemostatic processes. At equally efficacious antithrombotic dosages, TFV-1 caused neither severe thrombocytopenia nor bleeding in FcγRIIa-transgenic mice. Likewise, it did not induce hypocoagulation in human whole blood in the rotational thromboelastometry (ROTEM) assay used in perioperative situations. In contrast, TFV-3 and eptifibatide exhibited all of these hemostatic effects. Thus, the αIIbß3 antagonist, TFV-1, efficaciously prevents arterial thrombosis without adversely affecting hemostasis.