Current Pharmaceutical Design

ISSN: 1381-6128

Current Pharmaceutical Design
Volume 13, Number 28, 2007

Contents

Pharmacological Impact of Snake Venom Compounds
Executive Editor: C. Marcinkiewicz


Editorial Pp. 2851-2852


KTS and RTS-Disintegrins: Anti-Angiogenic Viper Venom Peptides Specifically Targeting the α₁ β₁ Integrin Pp. 2853-2859
J.J. Calvete, C. Marcinkiewicz and L. Sanz
[Abstract]


Anti-Angiogenesis and RGD-Containing Snake Venom Disintegrins Pp. 2860-2871
S. Swenson, S. Ramu and F.S. Markland
[Abstract]


Snake Venom Components Affecting Blood Coagulation and the Vascular System: Structural Similarities and Marked Diversity Pp. 2872-2886
Y. Yamazaki and T. Morita
[Abstract]


Snake Venom Proteins Affecting Platelets and Their Applications to Anti-Thrombotic Research Pp. 2887-2892
K.J. Clemetson, Q. Lu and J.M. Clemetson
[Abstract]


Importance of Snake Venom Metalloproteinases in Cell Biology: Effects on Platelets, Inflammatory and Endothelial Cells Pp. 2893-2905
A.M. Moura-da-Silva, D. Butera and I. Tanjoni
[Abstract]


Non-Lethal Polypeptide Components in Cobra Venom Pp. 2906-2915
Y.N. Utkin and A.V. Osipov
[Abstract]


Humanized Cobra Venom Factor: Experimental Therapeutics for Targeted Complement Activation and Complement Depletion Pp. 2916-2926
C-W. Vogel and D.C. Fritzinger
[Abstract]


Approaching the Golden Age of Natural Product Pharmaceuticals from Venom Libraries: An Overview of Toxins and Toxin Derivatives Currently Involved in Therapeutic or Diagnostic Applications Pp. 2927-2934
J.W. Fox and S.M.T. Serrano
[Abstract]


Trends in Snakebite Envenomation Therapy: Scientific, Technological and Public Health Considerations Pp. 2935-2950
J.M. Gutiérrez, B. Lomonte, G. León, A. Rucavado, F. Chaves and Y. Angulo
[Abstract]




Abstracts


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Editorial: Pharmacological Impact of Snake Venom Compounds

The snake venoms are the natural sources of many compounds that fascinated people through the history of pharmacology. Thus, the major part of logos of many national pharmaceutical organizations contains the image of a snake. Current drug design research considers venoms of many snakes as a valuable source for identification of new compounds with potential application in the pharmacology of many diseases. Moreover, toxic and non toxic components of snake venoms are very useful reagents for conducting medical research and understanding the physiology and pathology of many processes occurring in the body. Good examples are the snake venom disintegrins that inspired researchers to discover and characterize mammalian ADAM family proteins. The same snake venom disintegrins became a paradigm for the synthesis of new anti-platelet peptides and peptidomimetics that are currently used as therapy for certain heart and cardiovascular diseases. In this issue authors will describe the current stage and the future of research related to the compounds isolated from snake venoms. Among the many others, these compounds include for example anti-adhesive proteins such as disintegrins; enzymes such as phospholipases, metalloproteinases or serine proteases; and many neurotoxins affecting the central and peripheral nervous system. They are utilized in the pharmaceutical research of many diseases including blood coagulation, vasculature, nervous system, cancer, inflammatory diseases.

The first two review papers of this issue are devoted to snake venom disintegrins, in the context of their application in angiogenesis research. The first review [1] summarizes current knowledge about disintegrins that inhibit α1β1 integrin. This integrin is highly expressed on all types of microvascular endothelial cells and its participation in the neovascularization process has been established. The authors discuss structure-function correlation of the novel group of snake venom disintegrins that contain KTS or RTS motifs in their integrin-binding loop. These disintegrins are currently in the laboratory trials as a therapeutic approach toward inhibition of tumor vascularization.

Swenson and his colleagues [2] focused their review on the characterization of the effect of RGD-containing disintegrins on angiogenesis in various cancer models such as breast, ovarian and prostate. The RGD-disintegrins are the most investigated snake venom proteins, and in the past based on their structure two drugs, triofiban and eptifibatide were introduced for the therapy of certain cardiovascular diseases. The major interest of the authors is the homodimeric disintegrin contortrostatin and its inhibitory effect on the αvβ3 integrin present on endothelial cells. This recombinant disintegrin in a monomeric form appears to be a promising compound, and is an alternative to conventional anti-angiogenic agents.

The majority of viper venoms work on the prey by affecting blood coagulation and the vascular system. Many laboratories focused on these properties of venoms and isolated several compounds that are useful in pharmaceutical research related to these systems. The next two articles will characterize these compounds. Yamazaki and Morita [3] described a variety of toxins present in viper venoms that have unique biological activities that are not seen in mammals. These toxins affect platelet aggregation, blood coagulation cascade and vascular endothelial cells and have potentials in drug design. In this context, the authors also considered toxins from other organism including leeches and ticks.

The more oriented review on snake venom agents that affect hemostasis is presented by Clemetson et al. [4]. The aspect of using serine proteases, metalloproteases, C-type lectins, disintegrins and phospholipases is carefully discussed. The authors excellently summarized the effect of these compounds on platelets including investigation of receptor functions and signal transductions. It is interesting that properties of snake venom proteins lead to identification of new platelet receptors such as GPVI. Further studies of these proteins may provide new directions in diagnosis and therapy of human blood diseases.

The excellent review about particular group of snake venom proteins, metalloproteinases, is provided by Moura-da-Silve and her colleagues [5]. These enzymes are present in most viper venoms, playing a destructive role during the snakebite. These proteins are effective tools in the research of platelet function, angiogenesis, and pro-inflammatory reactions. Moreover, they are very useful in investigation of the mechanism of action of endogenous metalloproteinases in mammalian systems. It is intriguing that snake venom metalloproteinases have been characterized as toxins with catalytic and anti-adhesive properties.
The next two papers describe a pharmaceutical relation of peptides and proteins isolated from the cobra venom. Utkin and Osipov [6] summarize properties of non-lethal compounds of cobra venom that they classified into enzymes and non-enzymatic proteins. The varieties of enzymes are carefully characterized according to their ability to affect human systems including blood coagulation and complement. On the other hand, non-enzymatic proteins are extensively presented with special attention to three fingered toxins, nerve growth factors and protease inhibitors.

Vogel and Fritzinger [7] highly focused their article on one protein, cobra venom factor (CVF). This protein is a structural and functional analog of complement component C3. The authors discuss the possibility of application of CVF to inhibit complement in a clinical setting. CVF is a potent experimental tool for studying the biological function of complement and its involvement in pathology. They present an interesting idea of “humanization” of CVF in different forms that may be useful biopharmaceuticals for complement depletion in different clinical settings.

The excellent summary of the toxin-derived pharmaceuticals in therapy and diagnosis of human diseases are presented by Fox and Serrano [8]. The authors provide a clear overview of drugs currently accepted by the FDA, and those that are in clinical and pre-clinical trials. Inclusion in this review not only of snake, but all venous animals offers a much wider spectrum of the overview of pharmaceuticals that were designed on natural products. All of these products are summarized in three tables that will be helpful for readers to get a quick overview. Presentation of the web pages addresses allows easy access to the details describing a particular drug.

The last review is dedicated to a little different topic. Gutierrez and his colleagues [9] present articles about new directions in saving a human life after snakebite. This problem is a relevant public health issue in many regions of the world including subtropical countries of Africa, Asia, South America, and Oceania. Although the introduction of antivenoms significantly reduced the mortality of affected people, this therapy is ineffective in the neutralization of toxins responsible for local tissue damage. The authors overview new approaches to improve the quality and accessibility of antivenoms, and research stages for understanding and planning a therapy for neutralization of locally active toxins.

References

[1] Calvete JJ, Marcinkiewicz C, Sanz L. KTS and RTS-Disintegrins: Anti-Angiogenic Viper Venom Peptides Specifically Targeting the α₁ β₁ Integrin. Curr Pharm Des 2007; 13(28): 2853-2859.

[2] Swenson S, Ramu S, Markland FS. Anti-Angiogenesis and RGD-Containing Snake Venom Disintegrins. Curr Pharm Des 2007; 13(28): 2860-2871.

[3] Yamazaki Y, Morita T. Snake Venom Components Affecting Blood Coagulation and the Vascular System: Structural Similarities and Marked Diversity. Curr Pharm Des 2007; 13(28): 2872-2886.

[4] Clemetson KJ, Lu Q, Clemetson JM. Snake Venom Proteins Affecting Platelets and Their Applications to Anti-Thrombotic Research. Curr Pharm Des 2007; 13(28): 2887-2892.

[5] Moura-da-Silva AM, Butera D, Tanjoni I. Importance of Snake Venom Metalloproteinases in Cell Biology: Effects on Platelets, Inflammatory and Endothelial Cells. Curr Pharm Des 2007; 13(28): 2893-2905.

[6] Utkin YN, Osipov AV. Non-Lethal Polypeptide Components in Cobra Venom. Curr Pharm Des 2007; 13(28): 2906-2915.

[7] Vogel C-W, Fritzinger DC. Humanized Cobra Venom Factor: Experimental Therapeutics for Targeted Complement Activation and Complement Depletion. Curr Pharm Des 2007; 13(28): 2916-2926.

[8] Fox JW, Serrano SMT. Approaching the Golden Age of Natural Product Pharmaceuticals from Venom Libraries: An Overview of Toxins and Toxin-Derivatives Currently Involved in Therapeutic or Diagnostic Applications. Curr Pharm Des 2007; 13(28): 2927-2934.

[9] Gutiérrez JM, Lomonte B, León G, Rucavado A, Chaves F, Angulo Y. Trends in Snakebite Envenomation Therapy: Scientific, Technological and Public Health Considerations. Curr Pharm Des 2007; 13(28): 2935-2950.

Cezary Marcinkiewicz Ph.D.
Temple University, School of Medicine
Department of Neuroscience
Center for Neurovirology
1900 N.12th Street
Philadelphia, PA 19122
USA


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KTS and RTS-Disintegrins: Anti-Angiogenic Viper Venom Peptides Specifically Targeting the α₁ β₁ Integrin
J.J. Calvete, C. Marcinkiewicz and L. Sanz

Integrins α₁ β₁ and α₂ β₁ are highly expressed on the microvascular endothelial cells, and blocking of their adhesive properties significantly reduced the VEGF-driven neovascularization ratio and tumor growth in animal models. Hence, inhibitors of the α₁ β₁ and α₂ β₁ integrins, alone or in combination with antagonists of other integrins involved in angiogenesis (eg. α_v β₃, α_v β₅, and α₆ β₄), may prove benefitial in the control of tumor neovascularization. Viperidae snakes have developed in their venoms an efficient arsenal of integrin receptor antagonists. KTS- (obtustatin, viperistatin, lebestatin) and RTS- (jerdostatin) disintegrins represent viper venom peptides that specifically block the interaction of the α₁ β₁ integrin with collagens IV and I in vitro and angiogenesis in vivo. The possible therapeutic approach towards tumor neovascularization by targeting the α₅ β₁, α_v β₅ and α_v β₃ integrins with RGD-bearing disintegrins has been explored in a number of laboratories. Here we discuss structure-function correlations of the novel group of specific (K/R)TS-disintegrins targeting the α₁ β₁ integrin.


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Anti-Angiogenesis and RGD-Containing Snake Venom Disintegrins
S. Swenson, S. Ramu and F.S. Markland

Angiogenesis is the fundamental process by which new blood vessels are formed. Extensive research has shown that this event can be co-opted by tumors to ensure their growth, survival and metastasis. The study of tumor angiogenesis therefore represents a promising area of research for development of anti-cancer therapeutics. Integrins, a family of cell surface molecules, are a major target of interest as they are known to play a vital role in pathological angiogenesis. Remarkably, small disulfide-rich peptides known as disintegrins, isolated from the venoms of various snake species have been found to bind integrins with extremely high affinity and block their function. Disintegrins are capable of inhibiting several aspects of tumor cell behavior both in vitro and in vivo, including adhesion, migration, invasion, metastasis and angiogenesis. In this review, we will briefly discuss tumor angiogenesis and molecules implicated in the angiogenic process, with a special focus on the role of integrins. We will also discuss therapeutic approaches towards the treatment of tumor angiogenesis, including non-integrin-targeted agents currently in clinical trials. We will summarize the major findings from studies using disintegrins to target integrin-associated angiogenesis in cancer models. Finally, we will present results obtained in our laboratory using the novel dimeric disintegrin, contortrostatin (CN), in studies of endothelial cells and models of breast, ovarian and prostate cancer. In summary, disintegrins represent an exciting new class of molecules that can potentially be used in a clinical setting to inhibit angio-genesis and augment conventional chemotherapeutic agents in the treatment of cancer.


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Snake Venom Components Affecting Blood Coagulation and the Vascular System: Structural Similarities and Marked Diversity
Y. Yamazaki and T. Morita

In studies of blood coagulation and the vascular system, snake venom toxins have been indispensable in elucidating the complex physiological mechanisms that govern coagulation and the vascular system in mammals, given their potency and highly specific biological effects. The various components of snake venom toxins can be classified according to their mechanism of action, for example, serine proteases, metalloproteinases, Kunitz-type protease inhibitors, phospholipases A_{2, L}-amino acid oxidases, C-type lectin(-like) proteins, disintegrins, vascular endothelial growth factors, three-finger toxins, and cysteine-rich secretory proteins. Although the molecular structures of most toxins are not unique to snake venom toxins, venom proteins often exhibit marked diversity in their biological effects, despite their structural similarities. In this review, we identify several snake venom toxins capable of affecting blood coagulation and the vascular system, as well as various toxins from other organisms, including leeches and ticks.


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Snake Venom Proteins Affecting Platelets and Their Applications to Anti-Thrombotic Research
K.J. Clemetson, Q. Lu and J.M. Clemetson

Snake venoms are very complex mixtures of biologically active proteins and peptides that may affect hemostasis in many ways, by activating or inhibiting coagulant factors or platelets, or by disrupting endothelium. They have been classified into various families, including serine proteases, metalloproteinases, C-type lectins, disintegrins and phospholipases. The various members of a particular family act selectively on different blood coagulation factors, blood cells or tissues. Venom proteins affect platelet function in particular by binding to and blocking or clustering and activating receptors or by cleaving receptors or von Willebrand factor. They may also activate protease-activated receptors or modulate ADP release or thromboxane A₂ formation. L-amino acid oxidases activate platelets by producing H₂O₂. Many of these purified components are valuable tools in platelet research, providing new information about receptor function and signaling.


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Importance of Snake Venom Metalloproteinases in Cell Biology: Effects on Platelets, Inflammatory and Endothelial Cells
A.M. Moura-da-Silva, D. Butera and I. Tanjoni

Snake venom metalloproteinases (SVMPs) are widely distributed in snake venoms and play important roles in hemostatic disorders and local tissue damage that follows snakebite. The impact of SVMPs on hemostasis has been extensively studied showing diverse effects both on soluble factors and cellular components. The action of SVMPs involves catalytic and anti-adhesive properties, as well as direct cellular activation and/or the release of endogenous bioactive components. The purpose of this review is to overview the action of SVMPs on the inhibition of platelet functions; angiogenesis, particularly inducing apoptosis of endothelial cells; and regarding the pro-inflammatory reaction that follows snakebite. We discuss the structural features of the molecules that may be involved in such activities. The versatility and availability of SVMPs make them important tools for cell biology research into the mechanisms of action of endogenous metalloproteinases, for insights into cellular-matrix interactions and for clinical investigations into the treatment of snakebites.


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Non-Lethal Polypeptide Components in Cobra Venom
Y.N. Utkin and A.V. Osipov

Snakes from several genera (mostly from Naja genus) belonging to the Elapidae family are usually named cobras. The effect of cobra bites is mainly neurotoxic. This is explained by the presence of highly potent α-neurotoxin in their venoms. The other two highly toxic components of cobra venoms are cytotoxins and phospholipases A₂. These three types of toxins constitute a major part of cobra venom. They have attracted the attention of researchers for many years and have been very well studied and thoroughly described. However cobra venoms contain also many other less abundant components which possess very low toxicity or even are not toxic at all. These components, mostly proteins, belong to different structural and functional types, and the reason for their presence in the venom is not always evident. Some of them are known for many years (e.g., nerve growth factor and cobra venom factor); others (e.g., cysteine rich secretory proteins, CRISPs) were discovered only recently. There are non-lethal proteins with unique biological activities that can be used as biochemical tools, while others may be regarded as potential leads for drug design. This review is the first attempt to systemize the available data on non-lethal components of cobra venom.


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Humanized Cobra Venom Factor: Experimental Therapeutics for Targeted Complement Activation and Complement Depletion
C-W. Vogel and D.C. Fritzinger

Cobra Venom Factor (CVF) is the complement-activating protein in cobra venom. CVF is a structural and functional analog of complement component C3. In serum, CVF forms a physicochemically stable and control-resistant C3/C5 convertase that continuously activates C3 and C5, ultimately leading to depletion of serum complement. As CVF can be safely administered to vertebrate animals, it has become an important tool for complement depletion to study the biological functions of complement and its role in the pathogenesis of disease. CVF has also been used for targeted complement activation by chemically coupling it to monoclonal antibodies. Complement depletion is an attractive concept for pharmacological intervention in diseases where complement activation is part of the pathogenetic mechanism. Toward that end, the structural homology of CVF and C3 has been exploited by creating hybrid proteins in which short portions of C3 sequence have been exchanged with corresponding portions of CVF that introduce the desired ability of forming a stable convertase into human C3. These human C3 derivatives are "humanized CVF" proteins that represent an attractive biopharmaceutical for therapeutic complement depletion.


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Approaching the Golden Age of Natural Product Pharmaceuticals from Venom Libraries: An Overview of Toxins and Toxin Derivatives Currently Involved in Therapeutic or Diagnostic Applications
J.W. Fox and S.M.T. Serrano

Poisons and the toxins found in venomous and poisonous organisms have been the focus of much research over the past 70 years, most of which has been directed at understanding the biochemical and physiological mechanisms by which they elicit their dramatic pathological consequences. Much knowledge has been gained in terms of how poisons and venoms and their composite toxins give rise to the syndromes associated with envenoming and poisoning and in some isolated cases there have been a few such agents promoted for therapeutic use. However, it has only been in the past decade that an explosion of interest has occurred in mining these natural, highly evolved libraries of bioactive toxins and poisons for use in pharmacotherapeutics as drugs or drug leads as well as in diagnostic applications. We ascribe this recent phenomenon to advances in toxinology which have provided investigators with a relatively thorough understanding of the nature of venoms and their biologically active toxins: particularly with regard to the peptidomes and proteomes of venoms. This is in conjunction with our greatly improved understanding of the etiology of many human diseases and the identification of sites of potential therapeutic intervention. In this review we provide an overview of some of the toxins, toxin derivatives or poisons from animal venoms and secretions which are in various stages of development for use as pharmaceuticals or diagnostics in human diseases. As one will recognize, developments in this field suggest that toxinology is now entering a golden age in terms of the identification and use of toxins as potent novel pharmaceuticals.


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Trends in Snakebite Envenomation Therapy: Scientific, Technological and Public Health Considerations
J.M. Gutiérrez, B. Lomonte, G. León, A. Rucavado, F. Chaves and Y. Angulo

The therapy of snakebite envenomation has been based on the parenteral administration of animal-derived antivenoms. Despite the success of this treatment at reducing the impact of snakebite mortality and morbidity, mostly due to their capacity to neutralize systemically-acting toxins, antivenoms are of relatively low efficacy in the prevention of venom-induced local tissue damage, which often leads to permanent disability. The issue of safety also remains a concern, particularly for some antivenoms which induce a relatively high incidence of adverse reactions. Consequently, there is a need to improve the therapy of snakebite envenomations on the following lines: (a) the technologies to produce antivenoms require improvements aimed at obtaining more refined preparations of higher efficacy and safety, while being affordable for the public health systems of developing countries. (b) The growing knowledge on the biochemistry and toxicology of snake venoms should pave the way for the identification of natural and synthetic inhibitors of venom toxins, particularly of those involved in local tissue pathology. Such inhibitors might become a highly effective therapeutic tool for the abrogation of venom-induced local tissue damage. (c) A better knowledge of the inflammatory events secondary to venom actions may open the possibility of modulating such response, in order to prevent further tissue damage and to promote successful tissue repair and regeneration. A global partnership, involving many participants and combining scientific, technological and public health actions, is required to achieve a leap forward in the treatment of snakebite envenomations world-wide.