title: Evaluation of homogeneity and genetic stability of REOLYSIN (pelareorep) by complete genome sequencing of reovirus after large scale production.
abstract: REOLYSIN (pelareorep) is a proprietary isolate of the reovirus T3D (Type 3 Dearing) strain which is currently being tested in clinical trials as an anticancer therapeutic agent. Reovirus genomes are composed of ten segments of double-stranded ribonucleic acid (RNA) characterized by genome size: large (L1, L2, and L3), medium (M1, M2, and M3), and small (S1, S2, S3, and S4). The objective of this work was to evaluate the homogeneity and genetic stability of REOLYSIN. Sanger sequencing (SS) performed on test articles derived from the Master Virus Bank (MVB) and Working Virus Bank (WVB) identified many modifications when compared to GenBank reference sequences. Massively parallel sequencing (MPS) using Roche-454 sequencing was performed on REOLYSIN (100 L scale) and resulted in 69,821,115 bases and an average of 335 bases per read. Twenty-nine high confidence differences relative to the GenBank reference sequence were identified in REOLYSIN by MPS. Of those, 27 were previously identified by SS in the virus bank-derived test articles. Of the remaining two nucleotide differences, one was predicted to be silent at the amino acid level (L3 genome-T3163C, codon 1054, 86% of the population was "T" and 13% of the population were reported as "C"). The other modification was in the noncoding region (M1 genome-A2284A to A2284G), and A2284G was present in 97% of the population. The results obtained from MPS were comparable to those from SS; both demonstrate a high level of homogeneity at the amino acid level and genetic stability of REOLYSIN. Finally, phylogenetic analysis of the REOLYSIN L1 genome segment showed close evolutionary relationship with its human homologs, serotypes Lang and Dearing.
authors.affiliation: Oncolytics Biotech Inc., 210, 1167 Kensington Cr. NW, Calgary, AB, Canada, T2N 1X7.
authors.forename: Romit
authors.initials: R
authors.lastname: Chakrabarty
authors.forename: Hue
authors.initials: H
authors.lastname: Tran
authors.forename: Yves
authors.initials: Y
authors.lastname: Fortin
authors.forename: Zhenbao
authors.initials: Z
authors.lastname: Yu
authors.forename: Shi-Hsiang
authors.initials: SH
authors.lastname: Shen
authors.forename: John
authors.initials: J
authors.lastname: Kolman
authors.forename: David
authors.initials: D
authors.lastname: Onions
authors.forename: Robert
authors.initials: R
authors.lastname: Voyer
authors.forename: Allison
authors.initials: A
authors.lastname: Hagerman
authors.forename: Sarah
authors.initials: S
authors.lastname: Serl
authors.forename: Amine
authors.initials: A
authors.lastname: Kamen
authors.forename: Brad
authors.initials: B
authors.lastname: Thompson
authors.forename: Matt
authors.initials: M
authors.lastname: Coffey
mesh_terms: D016679:Genome, Viral / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D010802:Phylogeny; D012087:Reoviridae / Q000145:classification / Q000235:genetics*