Effect of the incorporation of hydroxy-terminated liquid silicones on the cure characteristics, morphology, and release of a model protein from silicone elastomer-covered rods

Silicone elastomer systems have previously been shown to offer potential for the sustained release of protein therapeutics. However, the general requirement for the incorporation of large amounts of release enhancing solid excipients to achieve therapeutically effective release rates from these otherwise hydrophobic polymer systems can detrimentally affect the viscosity of the precure silicone elastomer mixture and its curing characteristics. The increase in viscosity necessitates the use of higher operating pressures in manufacture, resulting in higher shear stresses that are often detrimental to the structural integrity of the incorporated protein. The addition of liquid silicones increases the initial tan delta value and the tan delta values in the early stages of curing by increasing the liquid character (G '') of the silicone elastomer system and reducing its elastic character (G'), thereby reducing the shear stress placed on the formulation during manufacture and minimizing the potential for protein degradation. However, SEM analysis has demonstrated that if the liquid character of the silicone elastomer is too high, the formulation will be unable to fill the mold during manufacture. This study demonstrates that incorporation of liquid hydroxy-terminated polydimethylsiloxanes into addition-cure silicone elastomer-covered rod formulations can both effectively lower the viscosity of the precured silicone elastomer and enhance the release rate of the model therapeutic protein bovine serum albumin. (C) 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011

Energy Dependence of Gold Nanoparticle Radiosensitization in Plasmid DNA

Gold nanoparticles (GNPs) are of considerable interest for use as a radiosensitizer, because of their biocompatibility and their ability to increase dose deposited because of their high mass energy absorption coefficient. Their sensitizing properties have been verified experimentally, but a discrepancy between the experimental results and theoretical predictions suggests that the sensitizing effect does not depend solely on gold's superior absorption of energetic photons. This work presents the results of three sets of experiments that independently mapped out the energy dependence of the radiosensitizing effects of GNPs on plasmid DNA suspended in water. Incident photon energy was varied from 11.8 to 80 keV through the use of monochromatic synchrotron and broadband X-rays. These results depart significantly from the theoretical predictions in two ways: First, the sensitization is significantly larger than would be predicted; second, it does not vary with energy as would be predicted from energy absorption coefficients. These results clearly demonstrate that the effects of GNP-enhanced therapies cannot be predicted by considering additional dose alone and that a greater understanding of the processes involved is necessary for the development of future therapeutics.

Structural basis for the inhibition of the essential Plasmodium falciparum M1 neutral aminopeptidase

Plasmodium falciparum parasites are responsible for the major global disease malaria, which results in > 2 million deaths each year. With the rise of drug-resistant malarial parasites, novel drug targets and lead compounds are urgently required for the development of new therapeutic strategies. Here, we address this important problem by targeting the malarial neutral aminopeptidases that are involved in the terminal stages of hemoglobin digestion and essential for the provision of amino acids used for parasite growth and development within the erythrocyte. We characterize the structure and substrate specificity of one such aminopeptidase, PfA-M1, a validated drug target. The X-ray crystal structure of PfA-M1 alone and in complex with the generic inhibitor, bestatin, and a phosphinate dipeptide analogue with potent in vitro and in vivo antimalarial activity, hPheP[CH2] Phe, reveals features within the protease active site that are critical to its function as an aminopeptidase and can be exploited for drug development. These results set the groundwork for the development of antimalarial therapeutics that target the neutral aminopeptidases of the parasite.

Balancing Inflammatory, Lipid, and Xenobiotic Signaling Pathways by VSL#3, a Biotherapeutic Agent, in the Treatment of Inflammatory Bowel Disease

Background: The interleukin 10 knockout mouse (IL10-KO) is a model of human inflammatory bowel disease (IBD) used to Study host microbial interactions and the action of potential therapeutics. Using Affymetrix data analysis, important signaling pathways and transcription factors relevant to gut inflammation and antiinflammatory probiotics were identified.

Methods: Affymetrix microarray analysis on both wildtype (WT) and IL10-KO mice orally administered with and without the probiotic VSL#3 was performed and the results validated by real-time polymerase chain reaction (PCR), immunocytochemistry, proteomics, and histopathology. Changes in metabolically active bacteria were assessed with denaturing gradient gel electrophoresis (DGGE).

Results: Inflammation in IL10-KO mice was characterized by differential regulation of inflammatory, nuclear receptor, lipid, and xenobiotic signaling pathways. Probiotic intervention resulted in downregulation of CXCL9 (fold change [FC] = -3.98, false discovery rate [FDR] = 0.019), CXCL10 (FC = -4.83, FDR = 0.0008), CCL5 (FC -3.47 FDR = 0.017), T-cell activation (Itgal [FC = -4.72, FDR = 0.00009], Itgae [FC = -2.54 FDR = 0.0044]) and the autophagy gene IRGM (FC = -1.94, FDR = 0.01), a recently identified susceptibility gene in human IBD. Consistent with a marked reduction in integrins, probiotic treatment decreased the number of CCL5+ CD3+ double-positive T Cells and upregulated galectin2, which triggers apoptosis of activated T cells. Importantly, genes associated with lipid and PPAR signaling (PPAR alpha [FC = 2.36, FDR = 0.043], PPARGC1 alpha [FC 2.58, FDR = 0.016], Nrld2 [FC = 3.11, FDR = 0.0067]) were also upregulated. Altered microbial diversity was noted in probiotic-treated mice.

Conclusions: Bioinformatics analysis revealed important immune response. phagocytic and inflammatory pathways dominated by elevation of T-helper cell 1 type (TH1) transcription factors in IL10-KO mice. Probiotic intervention resulted in a site-specific reduction of these pathways but importantly upregulated PPAR, xenobiotic, and lipid signaling genes. potential antagonists of NF-kappa B inflammatory pathways.

Inhibition of ribosome biogenesis induces apoptosis in p53-/- cells

The nucleolus is an important region of the nucleus that acts as the main site of rRNA transcription by RNA polymerase I (Pol-I), and one of the most prominent morphological markers of proliferative and invasive cancers is increased nucleolar size. Increases in Pol-I transcription leads to increased levels of ribosome biogenesis that are able to fuel rapid cell growth and proliferation due to increased ribosome numbers. Therefore Pol-I transcription seems a viable target for the development of anticancer therapeutics as abrogation of Pol-I transcription leads to cessation of cell growth and eventual cell death. We have confirmed that ellipticine compound, 9-Hydroxyellipticine (9HE), is an efficient inhibitor of Pol-I transcription in vitro and in p53+/+ and -/- cell lines in vivo. Short-term treatments (≤24 h) with micromolar concentrations of 9HE leads to decreased cell viability and proliferation, and leads to activation of caspases 3, 8 and 9, indicating that both intrinsic and extrinsic cell death mechanisms are activated upon Pol-I inhibition. Reactive oxygen species levels were also studied following short and long term treatments with 9HE and there was a 2/3-fold increase in cellular ROS levels. Long-term 9HE treatment (≥24 h) leads to cellular senescence as indicated by increased cellular morphology and senescence associated β-galactosidase staining, and this senescence is not accompanied by induction of autophagy. Following 24 h treatment there is also accumulation of cells in the G0/G1 phase of the cell cycle and qPCR analysis of cell cycle regulators shows down-regulation of G1/S transition associated cyclins. These data show that Pol-I transcription is a viable target for the development of novel chemotherapeutics, although further delineation of the cell death pathways remains. To further elucidate the mechanism, the role of mitochondrial death signals will be investigated by determination of cytochrome c release and regulation of Bcl2 family member proteins.

Fulfilling the 3Rs: Galleria mellonella as an In Vivo Model to Assess the Antimicrobial Efficacy of Self-assembling Peptide Hydrogels

The concept of ‘The Three Rs’ (The 3Rs: reduction, refinement and replacement) is an important consideration in the development of alternatives to animal testing in medical research. Invertebrate models such as Galleria mellonella are advantageous both economically and ethically.1 Galleria have proven to be effective alternatives to assess the antimicrobial activity of novel therapeutics.2
In this study Galleria mellonella are validated and used as an in vivo infection model to determine the antimicrobial activity of a novel self-assembling antimicrobial peptide NapFFKK.3 The peptide was considered as being non-toxic to the Galleria with 100% survival 120 hours post inoculation with NapFFKK. Following inoculation with Pseudomonas aeruginosa PAO1, Escherichia coli ATCC 11303, Staphylococcus epidermidis ATCC 35984 and Staphylococcus aureus ATCC 6538, the highest concentration allowing survival was selected and used as the test inoculum. Haemolymph was extracted from inoculated and peptide treated Galleria at either 24 or 72 hours post-treatment. Reduction in bacterial load was determined in comparison to a positive control. Bacterial load was decreased in all treated Galleria with decreasing antimicrobial activity demonstrated with a decreased concentration of peptide (2- log cycle reduction achieved in Escherichia coli inoculated Galleria treated with 2% NapFFKK). The results are promising regarding the use of Galleria mellonella as an infection model and NapFFKK as an effective novel antimicrobial.

Antibacterial Activity of Peptides from the African Volcano Frog

Introduction: Cationic, α- helical antimicrobial peptides found in skin secretions of the African Volcano Frog, Xenopus amieti include magainin-AM1, peptide glycine-leucine-amide (PGLa-AM1) and caerulein-precursor fragment (CPF-AM1). Objectives: The principle objective of this study was to determine the antibacterial activity of these peptides against a range of aerobic and anaerobic and oral pathogens. Secondary objectives were to establish their lipopolysaccharide (LPS) binding activity and determine potential cytotoxic effects against host cells. Methods: Magainin-AM1, PGLa-AM1 and CPF-AM1 were assessed for their antimicrobial activity against Fusobacteriim nucleatum, Streptococcus mutans, Lactobacillus acidophilus, Enterococcus faecalis and Streptococcus milleri using a double layer radial diffusion assay. The propensity for each peptide to bind LPS was determined using an indirect ELISA. The potential cytotoxicity of the peptides against human pulp cells in vitro was determined using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results: Magainin-AM1, PGLa-AM1 and CPF-AM1 displayed potent antimicrobial activity against all the bacterial pathogens tested, with Magainin-AM1 being the least effective. PGLa-AM1 was most potent against S. mutans, with a minimum inhibitory concentration (MIC) of 1.2 μM. PGLa-AM1 and CPF-AM1 were both very active against F. nucleatum with MIC values of 1.5 μM and 2.2 μM respectively. The LPS binding ability of the peptides varied depending on the bacterial source of the LPS, with PGLa-AM-1 being the most effective at binding LPS. Cytotoxicity studies revealed all three peptides lacked cytotoxic effects at the concentrations tested. Conclusions: The peptides magainin-AM1, PGLa-AM1 and CPF-AM1 from the African Volcano Frog, Xenopus amieti displayed potent antimicrobial activity and LPS binding activity against a range of oral pathogens with little cytotoxic effects. These peptides merit further studies for the development of novel therapeutics to combat common oral bacterial infections.

Development of human central nervous system 3D in vitro models for preclinical research

Neurological disorders are a major concern in modern societies, with increasing prevalence mainly related with the higher life expectancy. Most of the current available therapeutic options can only control and ameliorate the patients’ symptoms, often be-coming refractory over time. Therapeutic breakthroughs and advances have been hampered by the lack of accurate central nervous system (CNS) models. The develop-ment of these models allows the study of the disease onset/progression mechanisms and the preclinical evaluation of novel therapeutics. This has traditionally relied on genetically engineered animal models that often diverge considerably from the human phenotype (developmentally, anatomically and physiologically) and 2D in vitro cell models, which fail to recapitulate the characteristics of the target tissue (cell-cell and cell-matrix interactions, cell polarity). The in vitro recapitulation of CNS phenotypic and functional features requires the implementation of advanced culture strategies that enable to mimic the in vivo struc-tural and molecular complexity. Models based on differentiation of human neural stem cells (hNSC) in 3D cultures have great potential as complementary tools in preclinical research, bridging the gap between human clinical studies and animal models. This thesis aimed at the development of novel human 3D in vitro CNS models by integrat-ing agitation-based culture systems and a wide array of characterization tools. Neural differentiation of hNSC as 3D neurospheres was explored in Chapter 2. Here, it was demonstrated that human midbrain-derived neural progenitor cells from fetal origin (hmNPC) can generate complex tissue-like structures containing functional dopaminergic neurons, as well as astrocytes and oligodendrocytes. Chapter 3 focused on the development of cellular characterization assays for cell aggregates based on light-sheet fluorescence imaging systems, which resulted in increased spatial resolu-tion both for fixed samples or live imaging. The applicability of the developed human 3D cell model for preclinical research was explored in Chapter 4, evaluating the poten-tial of a viral vector candidate for gene therapy. The efficacy and safety of helper-dependent CAV-2 (hd-CAV-2) for gene delivery in human neurons was evaluated, demonstrating increased neuronal tropism, efficient transgene expression and minimal toxicity. The potential of human 3D in vitro CNS models to mimic brain functions was further addressed in Chapter 5. Exploring the use of 13C-labeled substrates and Nucle-ar Magnetic Resonance (NMR) spectroscopy tools, neural metabolic signatures were evaluated showing lineage-specific metabolic specialization and establishment of neu-ron-astrocytic shuttles upon differentiation. Chapter 6 focused on transferring the knowledge and strategies described in the previous chapters for the implementation of a scalable and robust process for the 3D differentiation of hNSC derived from human induced pluripotent stem cells (hiPSC). Here, software-controlled perfusion stirred-tank bioreactors were used as technological system to sustain cell aggregation and dif-ferentiation. The work developed in this thesis provides practical and versatile new in vitro ap-proaches to model the human brain. Furthermore, the culture strategies described herein can be further extended to other sources of neural phenotypes, including pa-tient-derived hiPSC. The combination of this 3D culture strategy with the implemented characterization methods represents a powerful complementary tool applicable in the drug discovery, toxicology and disease modeling.

Overexpression of mutant ataxin-3 in mouse cerebellum induces ataxia and cerebellar neuropathology.

Machado-Joseph disease (MJD), also known as spinocerebellar ataxia type 3 (SCA3), is a fatal, dominant neurodegenerative disorder caused by the polyglutamine-expanded protein ataxin-3. Clinical manifestations include cerebellar ataxia and pyramidal signs culminating in severe neuronal degeneration. Currently, there is no therapy able to modify disease progression. In the present study, we aimed at investigating one of the most severely affected brain regions in the disorder-the cerebellum-and the behavioral defects associated with the neuropathology in this region. For this purpose, we injected lentiviral vectors encoding full-length human mutant ataxin-3 in the mouse cerebellum of 3-week-old C57/BL6 mice. We show that circumscribed expression of human mutant ataxin-3 in the cerebellum mediates within a short time frame-6 weeks, the development of a behavioral phenotype including reduced motor coordination, wide-based ataxic gait, and hyperactivity. Furthermore, the expression of mutant ataxin-3 resulted in the accumulation of intranuclear inclusions, neuropathological abnormalities, and neuronal death. These data show that lentiviral-based expression of mutant ataxin-3 in the mouse cerebellum induces localized neuropathology, which is sufficient to generate a behavioral ataxic phenotype. Moreover, this approach provides a physiologically relevant, cost-effective and time-effective animal model to gain further insights into the pathogenesis of MJD and for the evaluation of experimental therapeutics of MJD.

Follow-up of three large community-based programs to reduce anaemia among children 24-59 months in Ghana, Malawi and Tanzania

L'anémie de l'enfant reste un problème d'importance pour la santé mondiale, malgré les décennies de recherche visant à comprendre son étiologie et à développer des interventions efficaces pour réduire sa prévalence et ses conséquences. Bien que les facteurs de risque individuels de l'anémie soient connus, y compris les facteurs liés à la malnutrition et à la morbidité, l'interaction entre lesdits facteurs est moins documentée dans des contextes où les enfants sont fréquemment exposés à plusieurs facteurs en même temps. Cette étude vise à documenter les efforts de lutte contre l'anémie du programme MICAH qui a été mis en oeuvre au Ghana, au Malawi et en Tanzanie. Ensuite, en utilisant les données relatives à la fois au processus et à l'évaluation colligées au cours du programme, elle vise à mieux comprendre les facteurs de risque d'anémie chez les jeunes enfants dans ces contextes et à comprendre comment les relations entre ces facteurs peuvent avoir changé au fil du temps lors de l'intervention. Spécifiquement, cette étude vérifie s‘il y a des preuves d'une réduction de la vulnérabilité des enfants aux facteurs de risque associés à l'anémie dans chaque contexte. Un examen de la documentation a été réalisé afin de caractériser le contexte du programme et des interventions, leur l'intensité et étendue. Les données transversales sur la nutrition et l'état de santé des enfants âgés de 24 à 59 mois (N = 2405) obtenues en 2000 et 2004 à partir des enquêtes d'évaluation du programme MICAH au Ghana, au Malawi et en Tanzanie, ont été utilisées pour décrire la prévalence de l'anémie. Les modèles polynomiaux de régression logistique et linéaire ont été utilisés pour estimer les risques d'anémie légère et d'anémie modérée / sévère et les niveaux d‘hémoglobine associés à des groupes de variables. Les estimations du risque attribuable à une population (RAP) ont aussi été calculées. Une anémie (Hb <110 g/L) a touché au moins 60% des enfants dans les trois pays; l'anémie modérée / sévère (<100 g/L) constituait la majorité des cas. Une forte diminution de l'anémie a été observée entre 2000 et 2004 au Ghana, mais seulement une légère baisse au Malawi et en Tanzanie. Le risque d'anémie modérée / sévère était associé au retard de croissance chez les enfants du Ghana (OR 2,68, IC 95% 1,70-4,23) et du Malawi (OR 1,71; 1,29-2,27) mais pas de la Tanzanie (OR 1,29; 0,87- 1,92). Le paludisme et les maladies récentes étaient associées à une hémoglobine plus basse. Une atténuation de cette association en 2004 a été observée seulement au Malawi pour le paludisme et au Ghana pour les maladies récentes. Le risque d'anémie modérée / sévère était 44% moindre chez les enfants âgés de 48 à 59 mois comparativement aux enfants de 24 à 35 mois dans les trois pays et cela n'a pas changé entre 2000 et 2004. Les RAP estimés ont montré qu‘environ un cinquième des cas d‘anémie modérée à sévère était attribuable au retard de croissance au Ghana et Malawi, mais pas en Tanzanie. Des RAP moindres et dépendants des contextes ont été trouvés pour le paludisme et les maladies récentes. Dans ces zones d‘intervention intégrées de santé et de nutrition la relation de certains facteurs de risque à l'anémie se modifia avec le temps. Le retard de croissance est resté toutefois un facteur de risque indépendant et non mitigé de l'anémie. Une réduction efficace des causes de la malnutrition chronique est nécessaire afin de réduire la vulnérabilité des enfants et de garantir un impact maximum des programmes de lutte contre l'anémie. Une mitigation de l'impact du paludisme peut par contre être visée dans les régions endémiques.

A study of the polycomb group complexes in the maintenance of heterochromatic genome stability and Alzheimer's disease.

La démence d'Alzheimer est une maladie neurodégénérative caractérisée par une perte progressive et irreversible des fonctions cognitives et des compétences intellectuelles. La maladie d’Alzheimer se présente sous deux formes: la forme familiale ou précoce (EOAD) qui représente 5% des cas et elle est liée à des mutations génétiques affectant le métabolisme des peptides amyloïde; et la forme tardive ou sporadique (LOAD) qui représente 95% des cas mais son étiologie est encore mal définie. Cependant, le vieillissement reste le principal facteur de risque pour développer LOAD. Les changements épigénétiques impliquant des modifications des histones jouent un rôle crucial dans les maladies neurodégénératives et le vieillissement lié à l'âge. Des données récentes ont décrit LOAD comme un désordre de l'épigénome et ont associé ce trouble à l'instabilité génomique. Les protéines Polycomb sont des modificateurs épigénétiques qui induisent le remodelage de la chromatine et la répression des gènes à l'hétérochromatine facultative. Nous rapportons que les souris hétérozygotes pour une protéine Polycomb développent avec l'âge un trouble neurologique ressemblant à LOAD caractérisé par l’altération des fonctions cognitives, la phosphorylation de la protéine tau, l'accumulation des peptides amyloïde, et le dysfonctionnement synaptique. Ce phénotype pathologique est précédé par la décondensation de l’hétérochromatine neuronale et l'activation de la réponse aux dommages à l'ADN. Parallèlement, une réduction d’expression de polycomb, malformations de l'hétérochromatine neuronale, et l'accumulation de dommages à l'ADN étaient également présents dans les cerveaux de patients LOAD. Remarquablement, les dommages de l'ADN ne sont pas distribués de façon aléatoire sur le génome mais sont enrichis au niveau des séquences répétitives. Les conclusions présentées dans cette thèse ont identifié des modifications épigénétiques spécifiques qui conduisent à une instabilité génomique aberrante menant à la formation de LOAD. Ces résultats vont aider au développement de nouveaux traitements qui peuvent potentiellement ralentir la neurodégénérescence.

Size effects in the magneto-optical response of Co nanoparticles

A study of the magneto-optical (MO) spectral response of Co nanoparticles embedded in MgO as a function of their size and concentration in the spectral range from 1.4 to 4.3 eV is presented. The nanoparticle layers were obtained by sputtering at different deposition temperatures. Transmission electron microscopy measurements show that the nanoparticles have a complex structure which consists of a crystalline core having a hexagonal close-packed structure and an amorphous crust. Using an effective-medium approximation we have obtained the MO constants of the Co nanoparticles. These MO constants are different from those of continuous Co layers and depend on the size of the crystalline core. We associate these changes with the size effect of the intraband contribution to the MO constants, related to a reduction of the relaxation time of the electrons into the nanoparticles.

Análisis del antagonismo entre el discurso del gobierno de Álvaro Uribe Vélez y el discurso de las comunidades indígenas de la Sierra Nevada de Santa Marta, en torno a las fumigaciones con glifosato (2002 - 2006)

El discurso del gobierno de Álvaro Uribe Vélez en su primer periodo presidencial, y el discurso de las comunidades indígenas de la Sierra Nevada de Santa Marta en torno a las fumigaciones con glifosato resultan en una relación antagónica, la cual produce una deslegitimación de la identidad cultural necesaria para la supervivencia de las comunidades indígenas. El análisis de los elementos de dicho antagonismo permite entender cómo se articula el discurso reivindicativo de las comunidades indígenas y cómo se fortifica la lucha reivindicativa por la legitimación de la identidad cultural. El discurso reivindicativo unifica las demandas generadas por dichas comunidades en torno a las fumigaciones con glifosato, mientras la lucha reivindicativa por la legitimación pretende resolver estas demandas y sentar las bases de un modelo de accionar político regido a partir de la aplicación de la palabra y el pensamiento tradicional de los pueblos indígenas.

Evolutionary analysis of novel serine proteases in the venom gland transcriptome of Bitis gabonica rhinoceros

Background: Serine proteases are major components of viper venom and target various stages of the blood coagulation system in victims and prey. A better understanding of the diversity of serine proteases and other enzymes present in snake venom will help to understand how the complexity of snake venom has evolved and will aid the development of novel therapeutics for treating snake bites. Methodology and Principal Findings: Four serine protease-encoding genes from the venom gland transcriptome of Bitis gabonica rhinoceros were amplified and sequenced. Mass spectrometry suggests the four enzymes corresponding to these genes are present in the venom of B. g. rhinoceros. Two of the enzymes, rhinocerases 2 and 3 have substitutions to two of the serine protease catalytic triad residues and are thus unlikely to be catalytically active, though they may have evolved other toxic functions. The other two enzymes, rhinocerases 4 and 5, have classical serine protease catalytic triad residues and thus are likely to be catalytically active, however they have glycine rather than the more typical aspartic acid at the base of the primary specificity pocket (position 189). Based on a detailed analysis of these sequences we suggest that alternative splicing together with individual amino acid mutations may have been involved in their evolution. Changes within amino acid segments which were previously proposed to undergo accelerated change in venom serine proteases have also been observed. Conclusions and Significance: Our study provides further insight into the diversity of serine protease isoforms present within snake venom and discusses their possible functions and how they may have evolved. These multiple serine protease isoforms with different substrate specificities may enhance the envenomation effects and help the snake to adapt to new habitats and diets. Our findings have potential for helping the future development of improved therapeutics for snake bites.

Sequence and phylogenetic analysis of viper venom serine proteases

Snakebites are a major neglected tropical disease responsible for as many as 95000 deaths every year worldwide. Viper venom serine proteases disrupt haemostasis of prey and victims by affecting various stages of the blood coagulation system. A better understanding of their sequence, structure, function and phylogenetic relationships will improve the knowledge on the pathological conditions and aid in the development of novel therapeutics for treating snakebites. A large dataset for all available viper venom serine proteases was developed and analysed to study various features of these enzymes. Despite the large number of venom serine protease sequences available, only a small proportion of these have been functionally characterised. Although, they share some of the common features such as a C-terminal extension, GWG motif and disulphide linkages, they vary widely between each other in features such as isoelectric points, potential N-glycosylation sites and functional characteristics. Some of the serine proteases contain substitutions for one or more of the critical residues in catalytic triad or primary specificity pockets. Phylogenetic analysis clustered all the sequences in three major groups. The sequences with substitutions in catalytic triad or specificity pocket clustered together in separate groups. Our study provides the most complete information on viper venom serine proteases to date and improves the current knowledge on the sequence, structure, function and phylogenetic relationships of these enzymes. This collective analysis of venom serine proteases will help in understanding the complexity of envenomation and potential therapeutic avenues.