Defective Endogenous Retroviruslike Particles of Chinese Hamster Ovary Cells

Anderson, Kevin; Low, Mari-Anne L.; Lie, Yolanda; Lazar, Richard; Keller, G A; Dinowitz, Marshall

doi:10.1016/b978-0-7506-1103-9.50014-7

Cited by 18 publications

(21 citation statements)

References 8 publications

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“…It is also clear from our analysis that the contaminating particles, though not characterized in detail, are of several types. It is worth underlining that such contaminations are not a unique finding, as production of retroviral or retrovirus-like particles by hybridoma cells (3,49,51) and even hamster cells such as CHO cells (the most frequently used cell line for the production of recombinant antibodies) (1,30) Fig. 3A.…”

Section: Discussionmentioning

confidence: 99%

Monoclonal Antibody 667 Recognizes the Variable Region A Motif of the Ecotropic Retrovirus CasBrE Envelope Glycoprotein and Inhibits Env Binding to the Viral Receptor

Dreja

Gros

Villard

et al. 2003

J Virol

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Monoclonal antibody (MAb) 667 is a neutralizing mouse monoclonal antibody recognizing the envelope glycoprotein (Env) of the ecotropic neurotropic murine retrovirus CasBrE but not that of other murine retroviruses. Since 667 can be used for preclinical studies of antiviral gene therapy as well as for studying the early events of retroviral infection, we have cloned its cDNAs and molecularly characterized it in detail. Spot technique-based experiments showed that 667 recognizes a linear epitope of 12 amino acids located in the variable region A of the receptor binding domain. Alanine scanning experiments showed that six amino acids within the epitope are critical for MAb binding. One of them, D 57 , is not present in any other murine retroviral Env, which suggests a critical role for this residue in the selectivity of 667. MAb 667 heavy-and light-chain cDNAs were functionally characterized by transient transfection into Cos-7 cells. Enzyme-linked immunosorbent assays and Biacore studies showed that the specificities as well as the antigen-binding thermodynamic and kinetic properties of the recombinant 667 MAb (r667) produced by Cos-7 cells and those of the parental hybridoma-produced MAb (h667) were similar. However, h667 was shown to contain contaminating retroviral and/or retrovirus-like particles which interfere with both viral binding and neutralization experiments. These contaminants could successfully be removed by a stringent purification protocol. Importantly, this purified 667 could completely prevent retrovirus binding to target cells and was as efficient as the r667 MAb produced by transfected Cos-7 cells in neutralization assays. In conclusion, this study shows that the primary mechanism of virus neutralization by MAb 667 is the blocking of the retroviral receptor binding domain of CasBrE Env. In addition, the findings of this study constitute a warning against the direct use of hybridoma cell culture supernatants for studying the initial events of retroviral cell infection as well as for carrying out in vivo neutralization experiments and suggest that either recombinant antibodies or highly purified antibodies are preferable for these purposes.CasBrE is a simple murine ecotropic retrovirus which causes a spongiform encephalopathy primarily affecting the motor center of the brain and the spinal cord. The virus was originally isolated from wild mice (17), and its neurovirulence is determined by the envelope glycoprotein (Env) sequence (13,46,47). Due to the poor replication ability of the virus in the brain, long periods of incubation (3 to 6 months) after neonatal infection are required for the onset of neuropathology. However, when the Env of CasBrE is introduced into the neuroinvasive but nonneuropathogenic Friend murine leukemia virus (MLV) strain FB29 in place of its natural Env, the resulting virus, FrCas E , induces a rapidly progressing, noninflammatory spongiform neurodegenerative disease with an incubation period of only 2 weeks.Retroviral envelope glycoproteins are synthesized as precursors ...

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Section: Discussionmentioning

confidence: 99%

Monoclonal Antibody 667 Recognizes the Variable Region A Motif of the Ecotropic Retrovirus CasBrE Envelope Glycoprotein and Inhibits Env Binding to the Viral Receptor

Dreja

Gros

Villard

et al. 2003

J Virol

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“…Endogenous RVLPs, as detected by transmission electron microscopy (TEM), are present in essentially all mammalian cell lines (11, 12). RVLP levels of 10 3 −10 8 particles/mL have been observed in typical industrial fermentation harvests employing CHO cells (13–15). By comparison, typical commercial bioreactors commonly have cell densities on the order of 10 6 cells/mL.…”

Section: Introductionmentioning

confidence: 95%

Performance of a Novel Viresolve NFR Virus Filter

Brough¹,

Antoniou²,

Carter³

et al. 2002

Biotechnol. Prog.

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Mammalian cell-expressed therapeutic proteins are particularly vulnerable to contamination by endogenous retrovirus-like particles (RVLPs). The Viresolve NFR filter was designed to meet the critical requirement of manufacturing a safe and virus-free therapeutic by retaining RVLPs by a minimum of six log reduction value (LRV). The NFR designation refers to retrovirus removal in a normal flow format. To qualify the product, we tested two model viruses: the 78 nm diameter phi6 bacteriophage and the 80-110 nm diameter Xenotropic Murine Leukemia Virus (X-MuLV). Robust retention was demonstrated over a wide range of process parameters. Viresolve NFR filters also retain other model adventitious viruses including 70-85 nm diameter Reovirus 3 (Reo3), 70-90 nm diameter Adenovirus 2 (Ad2), and 53 nm diameter PR772 by >6 LRV. In addition to these model viruses, the filter retains >7 LRV of both the mycoplasma Acholeplasma laidlawii and the bacterium Brevundimonas diminuta. Protein passage is shown to be consistently high (95-100%) for a variety of therapeutic protein products, including monoclonal antibodies. Characterization of the filter in specific applications is made simple by availability of ultralow surface area (5 cm(2)) disks, which are shown to scale linearly to the manufacturing scale pleated-filters. Viresolve NFR filters provide consistent water permeability performance (34-37 LMH/psi) and show very little plugging for all feedstocks evaluated. The Viresolve NFR filter incorporates Retropore, a unique asymmetric polyethersulfone membrane, the surface of which has been modified to minimize protein binding.

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“…Overall, this pilot study indicates specific expression of retroviral-related antigens in the baboon ovary. Other investigators have shown the presence of budding type C and intracytoplasmic type A particles in Chinese hamster ovary cells (21). However extensive screening failed to detect any evidence of infectivity of these retroviral particles (21).…”

Section: Expression Of Erv In the Ovary And Ovarian Tumoursmentioning

confidence: 99%

“…Expression of other retroviral-like proteins and mRNA transcripts, such as HERV-K, have also been demonstrated in normal adult mice testis and was found restricted to the undifferentiated spermatocytes as well as in normal human tissues (19,20). Budding type C and intracytoplasmic type A particles IAPs have been detected in Chinese hamster ovary cells (21) but infectivity of these retroviral particles has not been demonstrated (21).…”

Section: Expression Of Erv In Reproductive Tissuesmentioning

confidence: 99%