ReplicationSelective Oncolytic Adenovirus ElRegion Mutants Virotberapy for
II. Attributes of Replication-Selective Adenoviruses for Cancer Treatment 332 III. Biology of Human Adenovirus 332 IV. Mechanisms of Adenovirus-Mediated Cell Killing 333 V. Approaches to Optimizing Tumor-Selective Adenovirus Replication 333 VI. E1A-CR2 Region Deletion Mutants 334 VII. El B 55-kDa Gene Deletion Mutant d 1520 335 VIII. Clinical Trial Results with Replication-Competent Adenoviruses in Cancer Patients 337 A. Clinical Trial Results with Wild-Type Adenovirus 337 B. A Novel Staged...
IX Results from Clinical Trials with cf1520 OnyxOl 5 or CI1042
No maximally tolerated dose or dose-limiting toxicities were identified at doses up to 2xl012 particles administered by intratumoral injection. Flulike symptoms and injection-site pain were the most common associated toxicities 48 . This safety is remarkable given the daily or even twice-daily dosing that was repeated every 1-3 weeks in the head and neck region or pancreas 49 , Intraperitoneal, intraarterial, and intravenous administration were also remarkably well tolerated in general....
VI Homologous Recombination with Circular Ad Vector Genome Plasmids
Another way to circumvent the need for single cutting sites is obviously to use a circular instead of a linear template for the homologous recombination. However, in that case the selection process will necessarily involve a step of counterselection against the parental Ad plasmid, rendering the selection process more complex than the simple recircularization to be selected for in the systems described above. Two different approaches where the viral backbone is used as a supercoiled circular...
III Hybrid Adenoviral Vector Systems
A number of hybrid adenoviral vector systems have been reported in the literature, combining the properties of RV, AAV, and EBV vectors, as well as elements of other Ad serotypes, to enhance the therapeutic efficacy of Ad vectors in vivo. The principal aim of these new hybrid vectors is to overcome the limitations of transient Ad vector retention in infected cells. In addition to the well-documented limitations of Ad vectors Table II , some initially perceived advantageous properties of Ad...
Info Qlz
other cells e.g., by IFNa, IFNf , macrophage activation e.g., by IFNy , neutrophil recruitment and activation e.g., by IL-8, macrophage inflammatory protein MIP -2 , monocyte recruitment e.g., by MCP-1 , stimulation of NK cell-mediated clearance IL-12, IL-18 , stimulation of adaptive immunity TNFa, IL-2, IL-12, IL-18, others , and suppression of acute inflammation e.g., by IL-10, TGFp Table I . These and other cytokines elicited during in vivo microbial infection, together, form part of the...
A Master Cell Bank
Testing of the cell banks used in adenovirus vector production is of two general types safety testing and characterization. Table I is an overview of the recommended characterizations. The safety testing is intended to demonstrate that the cell bank is free of any detectable microbial contamination including bacterial, fungal, and viral. Sterility testing is a universal requirement for biologies and is set forth in 21 CFR 610.12. Alternative sterility assays validated to be of equal sensitivity...
IV Formulation and Stability
Process development must determine product stability over a wide range of conditions. The time course of the purification must be scrutinized for excessive run times and delays because rapid processing favors high yields. A well-designed process includes holding points selected such that the viral Figure 7 The most useful method for analyzing both crude and pure samples of adenovirus is anion-exchange HPLC. Pure adenovirus elutes in a nearly symmetrical peak. The adenoviral peak from lysate...
Propagation of Adenoviral Vectors Use of PERC6 Cells
W. W. Nichols, R. Lardenoije, B. J. Ledwith, K. Brouwer, S. Manam, R. Vogels, D. Kaslow, D. Zuidgeest, A. J. Bett, L Chen, M. van der Kaaden, S. M. Galloway, R. B. Hill, S. V. Machotka, C. A. Anderson, J. lewis, D. Martinez, J. Lebron, C. Russo, D. Valerio, and A. Bout II. Cells Expressing El of Adenovirus 134 A. Transformation of Cells by El of Adenovirus 134 B. El-Expressing Cell Lines for Adenoviral Vector Production 135 III. PER.C6 Prevents RCA during Vector Production 136 B. PF.R.C6...
XII VA RNA Genes
Adenovirus encodes one or two VA virus-associated RNAs, depending on serotype, of about 160 nucleotides that are transcribed by host cell RNA polymerase III. VA-RNAi targets the protein kinase named PKR reviewed in 91 . PKR is activated by the presence of low levels of double-stranded RNA, a likely product from symmetrical transcription of the Ad genome. Upon activation, PKR phosphorylates and inactivates eukaryotic initiation factor 2 alpha eIF-2a , thus inhibiting protein synthesis in...
V Homologous Recombination with Linear Ad Vector Genome Plasmids
The work by Chartier et al. 60 showed that stable maintenance of plasmids containing the entire Ad viral genome is achieved through the separation of the viral ITRs by the bacterial plasmid backbone, confirming the observations made earlier by Hanahan and Gluzman 64 . This was accomplished by the insertion of the left and right end of the Ad genome in their normal orientation into a colEI-derived bacterial plasmid ppolyll 65 , using conventional cloning techniques. Such a plasmid linearized...
IV Conclusion 1
The establishment of hybrid Ad vectors incorporating the advantageous properties of other viruses greatly expands their therapeutic potential. In the early 1990s, after the initial decade of proof-of-concept for Ad-mediated gene therapy, the main focus was on limiting the immunogenicity of the vectors to enhance transgene expression. Further restricting the expression of the highly immunogenic late viral transcripts by E4 deletions or by complete deletion of all viral genes in the gutless...
II ARCAs for Prostate Cancer CV706 and CV787
A. Adenovirus Gene Expression and Regulation Members of the human Adenoviridae family were first cultured from the tonsils and adenoids of children in 1953 29 . They represent 51 different serotypes which are distinguishable by antibody reactivity to epitopes on the virion surface. Each serotype is assigned to one of five subgroups A-E . Adenovirus type 5 Ad5 , a member of Subgroup C, is associated with a self-limiting, febrile respiratory illness and ocular disease in humans infectious virus...
VI Clinical Development of CV706 and CV787
CV706 and CV787 are novel therapeutic agents with a novel mechanism of action. As of this writing, a phase I II clinical trial of CV706 for recurrent local prostate cancer has been completed and CV787 has entered a multicenter Phase I II clinical trial for metastatic, hormonal refractory prostate cancer. A brief summary for CV706 trial result and factors impacting clinical efficacy and safety are discussed. A. CV706 Phase I II Trial for Locally Recurrent Prostate Cancer A Phase I trial of CV706...
V Safety Considerations of PERC6
A. QC Testing of PER.C6 Cells for Use in the Manufacture of Biologicals and Vaccines The safety of vaccines and biologicals manufactured in continuous cell lines of animal or human origin is of paramount importance and must be ensured by the manufacturer through a program of quality control QC testing applied to the product before release for human administration. This QC testing is intended to i ensure the identity of the product, ii ensure the safety and sterility of the product by...
Testing of Adenoviral Vector Gene Transfer Products FDA Expectations
Steven R. Bauer, Anne M. Pilaro, and Karen D. Weiss II. Manufacturing Control and Product Characterization 616 A. Purity, Safety, and Potcncy 616 B. Regulation of Process as Well as Product 617 C. Current Good Manufacturing Practices 617 III. Development of Recommendations for the Manufacture and Characterization of Adenoviral Vectors 618 IV. Considerations in Manufacturing Adenoviral Vectors 620 A. Components and Characterization 620 A. Standard Operating Procedures 621 B. Quality Assurance...