adenoviral vectors – BioProcessing Journal

Defining Therapeutic Window for Viral Vectors: A Statistical Framework to Improve Consistency in Assigning Product Dose Values

NOTE: Page 7 of this article has been revised to correct an error in the original article which stated that “..the probability of failing a lot when it actually is not equivalent, becomes 1-2*α.” The correct equation, 1-α, has replaced the errant one and a single sentence following it was added to further clarify this concept.

ABSTRACT: Pre-clinical and clinical trials conducted to establish the minimum effective dose and the maximum tolerated dose of a viral vector assume that the assigned dose values are comparable across studies. Toxicity has been associated with high dose administration of both adenovirus and adeno-associated virus-based vectors, and increased attention must be paid to assays used to measure dose. High assay variability can be mitigated by replication and the reporting of a mean value for product lot release. The establishment of a dose specification and a testing strategy must take into account the risk of errant quality control decisions. This can be accomplished by linking assay qualification information to measurement uncertainty through a statistical framework. By adopting an equivalence approach, the risk of releasing lots with unacceptably high or low dose values is minimized by reducing measurement uncertainty. This article provides a worked-through example to introduce applicable statistical concepts and the equations necessary to facilitate their implementation in the field.

Risk Analysis and Management Viral Vectors

adeno-associated viral vectors adenoviral vectors assay qualification dosage dose-determining assay equivalence limits therapeutic window

Evaluation of Filtration Products in the Production of Adenovirus Candidates Used in Vaccine Production: Overview and Case Study

Adenoviral vectors (AAV’s) offer a promising new approach to vaccine development. They have the ability to be rapidly manipulated for bearing transgenic coding for specific antigenic proteins, efficiently infect a variety of mammalian cell types (including antigen-presenting cells) and induce a broad immune response against the target antigen in vaccine recipients. Furthermore, AAV’s offer an excellent safety profile, in that they can be engineered to be non-replicating in the vaccine recipient and they lack the molecular mechanism for integration into the host genome. AAV’s are highly amenable to scalable manufacturing processes such as the use of stirred tank bioreactors, high capacity filtration methods, and chromatographic purification procedures…

Biologics Production Viral Vectors

adenoviral vectors concentration diafiltration filtration lysate clarification vaccines

Manufacturing and Testing of a Multi-Clade Adenoviral Vector-Based Candidate Vaccine Against Human Immunodeficiency Virus

Despite 20 years of intensive research, the development of an effective vaccine to combat the worldwide AIDS pandemic remains an elusive goal. Currently, more than 40 million individuals are infected with human immunodeficiency virus (HIV) and there have been more than 25 million related deaths. Globally, the rate of new infections is alarming, with ten new infections occurring every minute. Ninety-five percent of these infections occur in the developing world. Several significant challenges face the development of an effective HIV vaccine…

Manufacturing Viral Vectors

adenoviral vectors HIV immunology multi-clade vaccine vaccines

Advanced Methods of Adenovirus Vector Production for Human Gene Therapy: Roller Bottles, Microcarriers, and Hollow Fibers

Various types of viral vectors are being employed extensively as gene therapeutics to treat cancer and genetic diseases. Among the viruses that have been produced for human clinical trials (i.e. retrovirus, adenovirus, poxvirus, adeno-associated virus, and herpesvirus vectors) adenoviruses exhibit the lowest pathogenicity yet still infect an extensive range of cell types with high efficiency. These key characteristics make recombinant adenoviruses efficient gene-delivery vehicles and excellent research tools. However, the time-consuming and complex processes of generation, amplification, purification, and quality testing associated with production of recombinant adenoviruses make it difficult for many researchers to utilize these vectors. This is particularly true with respect to cell culture optimization and the virus propagation protocols employed in vector production. In this regard, the development of innovative cell culture techniques has become vital for optimizing vector production for gene therapy…

Biologics Production Cell & Gene Therapy Viral Vectors

adenoviral vectors analysis cell culture enumeration propagation purification recombinant proteins

Defining a Detailed Approach to Using the Adenovirus Reference Material (ARM)

Through the tremendous efforts of the Adenovirus Reference Material Working Group (ARMWG), an adenovirus reference material (ARM) is now available from the American Type Culture Collection (ATCC). The history and progress of the ARM production and characterization has been presented at many meetings and published in numerous journal articles. Although general statements have been made regarding how the ARM should be used, there is no formal directive or specific set of instructions detailing its application in the field. The goals of this paper are (1) to briefly review the objectives for development and implementation of the ARM, (2) to describe a critical assumption necessary to meet those objectives, (3) to outline specific approaches for using the ARM, and (4) to highlight the need for a working group to address the issues raised in the process…

Viral Reference Materials

adenoviral vectors adenovirus infectious titer particle concentration replication-competent standards

Adenovirus Reference Material: Determination of Particle Concentrations Obtained by Orthogonal, Physical-Chemical Methods

With the continued progress of adenoviral vectors in gene therapy studies it is increasingly evident that a more formalized approach to the characterization and analysis of these viral vectors is urgently needed. Today, adenoviral vectors are beginning to be considered “well characterized biologics,” as shown by numerous publications describing sophisticated analytical approaches for recombinant adenovirus product candidates. Because the analytical definitions of adenoviral vectors currently lack comparison to a common standard, the problem for regulatory agencies is how to objectively evaluate safety in relation to the administered dose. This well-recognized need for an adenovirus standard has been addressed by a consortium of representatives from regulatory agencies, industry, and academic organizations — the Adenoviral Reference Material (ARM) Working Group. Its work has come to fruition in the recent public availability of the ARM, a purified wild type 5 adenovirus. Many aspects of the history, production, and characterization of the ARM have been published in detail…

Regulatory Viral Reference Materials Viral Vectors

adenoviral vectors analysis characterization particle concentration safety standards

Viral Gene Vectors

An astonishing range of viruses has provided building blocks for gene delivery systems, from the simple adeno-associated virus with a 5 kb genome to the complex poxviruses with 300 kb. This review focuses on non-replicating viral vectors that infect host cells just once, without producing infections virus. Viral vectors are generally characterized by several criteria, including their ability to integrate into the host genome, coding capacity, titer, toxicity, immunogenicity, host range, duration of gene expression, and transient or stable production systems. These are precisely the features that need to be carefully studied in the context of the application when deciding which vector to use…

Biologics Production Cell & Gene Therapy Viral Vectors

adeno-associated viral vectors adenoviral vectors gene delivery gene therapy non-replicating viral vectors vaccines

Adenovirus Type 5 (Ad5) Chromatographic Purification Process at the 20 L Scale

Recombinant adenovirus are attractive as vectors for gene therapy because: they exhibit wide tissue tropism and high transduction efficiency; adenovirus cultures can reach high specific titers (10^10 VP/mL), and; their use in the treatment of cancer and other serious diseases is valuable. A primary mode of adenovirus purification continues to be CsCl density gradient centrifugation…

Viral Vectors

adenoviral vectors adenovirus adenovirus serotype 5 chromatography purification transduction

Methods for Detection and Evaluation of Replication Competent Adenovirus (RCA)

In the past, researchers developing gene therapy applications used replication-defective human Adenovirus 5 (Ad5) as a vector for delivering DNA sequences, almost exclusively. Ad5 vectors are typically rendered replication defective by the deletion of E1a gene sequences. A complementing cell line containing the E1a gene makes it possible to produce Ad5 vectors in large scale. Of the various cell lines that have been constructed for the purpose of high-titer Ad5 production, HEK293 cells and PER.C6 cells are the most widely used…

Biologics Production Viral Vectors

adenoviral vectors adenovirus serotype 5 replication replication-competent

Chemical Cell Lysis for Large-Scale Adenoviral Vector Production

Production of non-enveloped viruses generally requires a cell lysis procedure to liberate mature particles trapped within their host cells. The standard bench-scale practice of using freeze/thaw cycles is simple and effective, but heat transfer limitations restrict the technique to relatively small applications. Here we show that a ten-minute treatment with a dilute mixture of polysorbate-80 and tri-butyl phosphate effectively liberates adenovirus from host cells…

Biologics Production Viral Vectors

adenoviral vectors adenovirus cell lysis scale-up

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