![]() ![]() 1,6 Thus, each vector can present unique challenges during downstream processing. 5 Pseudotyping can also influence RV/LV stability, which may be improved or made worse depending on the vector and pseudotype component. ![]() The instability of these particles can lead to a loss of yield – potentially up to 70% during the whole downstream purification process. The presence of a lipid envelope makes these vectors less stable. The downstream processing of RV/LV vectors presents its own unique challenges. 2 Despite the ongoing debate over a possible benefit from the presence of empty particles in AAV therapies, these are still considered a major contaminant 3,4 and health authorities encourage sponsors to set limits on the amount of empty or partially full vectors. AAV vectors can produce a significant portion of “empty” particles devoid of transgene at up to 95% to total particles. Cell lysis can generate significant quantities of host cell contaminants including DNA and protein that increase purification burden. For example, AAV also accumulated in the media and is partly released from cells via a lytic process, AAV viruses are generally harvested by cell lysis to improve yield. The differences in physical characteristics between various vectors presents a diverse set of purification challenges that result in a multiplicity of needed techniques and solutions for the industry. Pseudotyping with VSV-g is reported to improve stability to some extent 1 Differences and similarities in 4 common viruses used as viral vectors VirusĪ. Table 1 shows some similarities and differences of the 4 commonly used viruses used as vectors: adenovirus (AdV), adeno-associated virus (AAV), retrovirus (RV), and lentivirus (LV). ![]() Viral vectors vary greatly in physical characteristics depending on the type of virus, the serotype of the same virus, and can sometimes vary with different transgene inserts within the same vector (AAV). In addition to providing purity, purification processes need to meet production scale, which can be quite large for clinical trials and commercial applications.Īpproaches to vector purification exploit the physical characteristics of the viral particle such as size, surface charge, and hydrophobicity. Viral vector batches destined for clinical use must comply to increasing regulatory standards for impurities and contaminants as these can affect product safety and potency. The goal of downstream processing is to separate the viral vector from the various impurities produced during upstream processing and to get the virus into the appropriate state for formulation and administration to patients. Downstream Manufacturing of Gene Therapy Vectors ![]()
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