Gene therapy is a fast growing technology that offers treatments for genetic diseases. The method is based on introducing genetic material into a patient to replace the disease-causing gene, using a vector. This report examines the potential of some viral vectors for gene therapy, to give Bio-Works Technologies a recommendation on what the future market demands. Oncolytic viruses, vaccines and gene editing are not treated in the report as a delimitation.  Viral vectors have different biological properties and require different purification methods, making them suitable for different applications in gene therapy. In the purification of the viruses it can be challenging to obtain a high purity and large-scale manufacturing. One major drawback with most purification methods is that they are not specific to just one virus, which leads to contaminants in the solution and lower purity. The viral vectors handled in the report are the adenovirus, adeno-associated virus, gammaretrovirus, lentivirus, alpharetrovirus, foamy virus, herpes simplex virus and baculovirus. These were chosen as they are relevant vectors for gene therapy and stay within the scope of the report. Lentiviral vectors (LVs) and adeno-associated viral vectors (AAVs) will dominate the gene therapy field in the coming years. This is based on the information that the use of AAVs and LVs in clinical trials have increased in recent years, while the other vectors mentioned above have slightly decreased or show no apparent change. However, challenges still remain in the purification processes. Ligands used in affinity chromatography for purification of AAVs are effective at removing most contaminants, but cannot distinguish between empty and loaded capsids, which can induce immune response when used clinically. This is the main challenge when purifying AAVs. The empty capsids can be removed with ion exchange chromatography, which results in higher purity but also lower recovery. There is no specific purifying method for LVs, therefore a lentivirus-specific affinity ligand, such as an antibody ligand, would be beneficial for the purification and manufacturing procedure.  In addition to AAVs and LVs, baculoviral vectors and foamy viral vectors show great potential in a long-term perspective but they only have been researched in preclinical studies. Moreover, herpes simplex viral vectors and adenoviral vectors show potential in cancer treatments or as vaccines rather than in augmentation gene therapy.

By participating in the market a party buys and sells different types of contracts resulting in the collection of contracts growing. With a large collection of contracts come the hurdles of an increasing operational cost, a harder-to-manage order book, and an increase in counterparty risk. To combat these problems we set out to minimize the size and quantity of contracts by performing what is called a compression. We have looked into three different types of compression methods for interest rate swap contracts. One method is specialized for central clearing, Coupon Blending, and two methods for bilateral clearing, Closed Loops, and the Network Simplex Method. By using Monte Carlo Simulations, all three methods could be compared to one another to conclude the significant findings. The clear winner for centrally cleared contracts was Coupon Blending which could terminate over 92% of the contracts, and reduce the total absolute size of the contracts by over 75%. Network Simplex came in as a close second which could also reduce the total absolute size of the contracts by over 75% but only terminate 86%. Coupon Blending and Network Simplex, both had very similar accuracy in their compression. However, NetworkSimplex performed better at keeping the system’s total risk intact. For bilateral clearing, NetworkSimplex performed the best where the Closed Loops strategy was not an optimized approach. / Genom att delta i den finansiella marknaden köper och säljer en participant olika sorters kontrakt vilket resulterar i att samlingen av kontrakt växer. Med en ständigt växande samling av kontrakt skapas problem som, att kostnaden för hantering ökar, att orderbokens hantering blir svårare och en ökad risk för konkurs. För att undvika dessa problem kan man utföra kompression vilket är att försöka reducera kontrakten i antal och storlek. Vi har studerat tre olika typer av kompressionsstrategier för kompression av ränteswappar. Den första strategin är Coupon Blending som är specialiserad för central clearing medan de två andra, Closed Loops och Network Simplex Metoden är utvecklade för bilateral clearing. Genom att använda Monte Carlo Simuleringar på alla tre strategier kunde vi dra slutsatser kring deras egenskaper och effektivitet. Den bästa strategin var Coupon Blending som kunde terminera över 92% av alla kontrakt, och samtidigt reducera den totala absoluta storleken på kontrakten med 75%. Network Simplex presterade också bra och kunde reducera den totala absoluta storleken på kontrakten med 75% och terminera 86% av kontrakten. Coupon Blending och Network Simplex hade bägge en liknande noggrannhet, men Network Simplex var något bättre på att hålla systemets totala risk intakt. För bilateral clearing presterade Network Simplex bäst där Closed Loops strategin inte var tillräckligt optimerad.

The obligate intracellular bacterium Chlamydia trachomatis is the most common bacterial STD agent in the US. This bacterium has a unique biphasic developmental cycle in which the infectious elementary body (EB) infects a host mucosal epithelial cell and differentiates into the replicative form (the reticulate body or RB) within a modified vacuole called an inclusion. The RB later divides and develops back into an EB and is released, perpetuating the infectious cycle. When developing chlamydiae are exposed to unfavorable environmental conditions, they deviate from the normal developmental cycle into a non-infectious but viable state termed persistence. Previous data from our laboratory indicate that i) during C. trachomatis/HSV co-infection, the chlamydiae become persistent and ii) HSV gD interaction with host cell surface is sufficient to induce this response. During viral entry, HSV gD interacts with one of four host co-receptors, one of which is the host adhesion molecule nectin-1. Interestingly, Western blotting demonstrated that nectin-1 is significantly decreased in C. trachomatis-infected HeLa cells. Additional studies indicated that active C. trachomatis replication is required for nectin-1 down-regulation and nectin-1 is likely down-regulated post-translationally. CPAF, a chlamydia-secreted protease, is responsible for degrading several host proteins. Both in vivo experiments using CPAF-specific chemical inhibitors and cell-free cleavage assays using recombinant CPAF indicate that nectin-1 is degraded by CPAF in C. trachomatis-infected cells. Further studies suggest that nectin-1 is the most likely candidate involved in triggering HSV-induced chlamydial persistence. Co-infection experiments using nectin-1-specific HSV-1 mutants suggest that nectin-1 is, indeed, required for persistence induction. Additional studies in single co-receptor-expressing CHO cells demonstrate that, despite the fact that HSV-1 enters both HVEM- and nectin-1-expressing cells, viral co-infection reduces chlamydial infectivity only in the CHO-nectin-1 cell line. These data confirm that HSV/nectin-1 interaction is sufficient for chlamydial persistence induction. Although nectin-1 ligation is known to activate Cdc42, pull-down assays indicate that Cdc42 is not activated in co-infected HeLa cells. Taken together, these data suggest that: i) HSV gD-nectin-1 binding activates a novel host epithelial cell pathway that restricts chlamydial development and ii) the chlamydiae may degrade nectin-1 to evade this inhibitory host response.

Previous studies have established that ozone-oxygen mixtures can be used to inactivate a variety of microorganisms including bacteria, fungi and viruses. Ozone is a potent reactive oxygen species (ROS) that rapidly decays into a variety of additional short half-life ROS which have been shown to cause oxidative damage to biological molecules. I hypothesize that controlled ozone exposure and the subsequent generation of additional ROS would reduce viral infectivity by lipid and/or protein peroxidation. A proprietary ozone-oxygen delivery system was used to inactivate a series of enveloped [herpes simplex virus type-1 strain McIntyre (HSV-1), vaccinia strain Elstree (VAC), vesicular stomatitis virus strain Indiana (VSV), and influenza A strain (H1N1) A/WS/33] and non-enveloped [human adenovirus type2 (Ad2)] viruses. Plaque reduction and suspension-infection viral antigen assays were used to determine inactivation kinetics. After ozonation, HSV-1 and VSV lost up to 6 log10 infectious particles in 15 min, while VAC and influenza A lost up to 5 log10 in 40 min and 30 min, respectively. In comparison, the non-enveloped Ad2 lost up to 5 log10 in 60 min. Increasing amounts of serum supplementation in the ozone treated virus suspensions slowed the rate of inactivation in both enveloped and non-enveloped viruses, suggesting the protective effect of serum against ozone. Lipid peroxidation was determined through a chromogenic assay for malondialdehyde (MDA), a byproduct of peroxidation events. MDA concentrations were inversely correlated with virus infectivity, as MDA concentrations elevated with virus exposure time to ozone. Transmission electron microscopy images of Ad2, HSV-1, VAC and VSV confirmed the drastic morphological changes that resulted from ozone treatment. The ROS-mediated attack compromised the integrity of the lipid envelopes and protein shells of the viruses. These data suggest that a wide range of viruses can be inactivated through use of an innovative ozone delivery system, thus validating my hypothesis.

A multi-commodity flow problem consists of moving several commodities from their respective sources to their sinks through a network where each edge has different costs and capacity constraints. This paper explores different linear programming algorithms and their performance regarding finding an optimal solution for multi-commodity flow problems. By testing several of different network constraints, we examine which algorithms are most suitable for specific network and problem structures. Furthermore, we implement our own multi-commodity solver and compare its performance against state-of-the-art linear programming solvers. The results show that for the methods we tested it is difficult to discern which class of linear programming methods are optimal solvers for multi-commodity flow problems and that their performance depends on how the network and commodities are structured.

Experiments were carried out to determine if pretreatment of cells with N-Methyl-N'-Nitro-N-Nitrosoguanidine (a potent alkylating agent known to induce DNA repair) would affect Herpes Simplex Virus Replication. The data demonstrated a 1.5 fold increase in virus yield, a 2 fold increase in HSV specific TK activity and no change in HSV specific DNA polymerase activity in MNNG treated cultures. The effects of MNNG treatment on virus replication and enzyme expression are discussed. / Thesis / Master of Science (MSc)

No description available.

No description available.

No description available.

No description available.