Candidate Cell Substrates, Vaccine Production, and Transmissible Spongiform Encephalopathies

Pedro Piccardo

, Larisa Cervenakova, Irina Vasilyeva, Oksana Yakovleva, Igor Bacik, Juraj Cervenak, Carroll McKenzie, Lubica Kurillova, Luisa Gregori, Kitty Pomeroy, and David M. Asher

Author affiliations: University of Edinburgh, Easter Bush, UK, (P. Piccardo); Food and Drug Administration, Kensington, Maryland, USA (P. Piccardo, I. Bacik, J. Cervenak, L. Kurillova, L. Gregori, K. Pomeroy, D.M. Asher); Holland Laboratory American Red Cross, Rockville, Maryland, USA (L. Cervenakova, I. Vasilyeva, O. Yakovleva, C. McKenzie)

Abstract

Transmissible spongiform encephalopathy (TSE) agents have contaminated human tissue–derived medical products, human blood components, and animal vaccines. The objective of this study was to determine the potential susceptibility to infection of 5 cell lines used or proposed for manufacture of biological products, as well as other lines. Cell lines were exposed to the infectious agents of sporadic and variant Creutzfeldt-Jakob disease and bovine spongiform encephalopathy (BSE). Exposed cultures were tested for TSE-associated prion protein (PrP^TSE) and TSE infectivity by assay in rodents and nonhuman primates. No PrP^TSE or infectivity has been detected in any exposed cell line under study so far. Animals inoculated with BSE brain homogenate developed typical spongiform encephalopathy. In contrast, animals inoculated with cells exposed to the BSE agent remained asymptomatic. All cell lines we studied resisted infection with 3 TSE agents, including the BSE agent.

Transmissible spongiform encephalopathies (TSEs or prion diseases) are a heterogeneous group of fatal neurodegenerative diseases that affect animals and humans. TSEs can be sporadic, transmitted iatrogenically, or expressed as familial disorders. TSEs include scrapie in sheep and goats, bovine spongiform encephalopathy (BSE) in cattle, chronic wasting disease in cervid ruminants, and mink encephalopathy. In humans the most common TSEs are sporadic, familial, and variant Creutzfeldt-Jakob disease (sCJD, fCJD, and vCJD, respectively). TSEs are characterized by the accumulation in the central nervous system and, less often, in lymphoid tissues of TSE-associated prion protein (PrP^TSE), a conformational variant of a normal host cellular prion protein (PrP^C). PrP^C is a nonessential protein but, at least in mice and cows, must be expressed by animals susceptible to TSE infection. There is compelling evidence that the BSE agent has infected humans, causing vCJD. Most cases of vCJD are attributed to exposure to contaminated beef products (1–3). In addition, vCJD infections have been transmitted by transfusions of nonleukoreduced erythrocyte concentrates and by a human-derived coagulation factor (factor VIII) (4–6).

The conclusion that PrP^TSE is central to the pathogenesis of TSE is based on the temporal and anatomic correlations between accumulation of PrP^TSE and the development of pathologic changes in tissues of the central nervous system (1). However, TSEs can develop in the absence of detectable PrP^TSE and, conversely, PrP^TSE might accumulate without causing either clinical illness or the neuropathologic alterations typical of TSEs (i.e., a progressive fatal illness with spongiform degeneration of the brain) (7,8). In short, the molecular basis of TSE infection and the role of PrP^TSE (unquestionably important in pathogenesis of TSEs) are not yet entirely clear, and both remain key issues in TSE research (9,10). The standard assay for detecting a TSE agent remains bioassay in susceptible animals.

Many investigators once believed that TSE agents infected mainly, if not exclusively, cells of neuronal and lymphoid lineages. It has become clear, however, that the susceptibility of cells to infection with TSE agents cannot be reliably predicted either from their tissue of origin or level of expression of PrP^C (11,12). Studies showing that murine fibroblast cell lines are susceptible to infection with mouse-adapted scrapie agent (11,12) increased concern that nonneuronal cell substrates used to propagate viruses for vaccine production might become infected with a TSE agent contaminating some component of culture medium, especially bovine serum (13). The theoretical risk of contaminating vaccines or other biologic products prepared in culture cells with TSE agents from animal-derived materials in media has been considered low. However, 1) as noted above, the blood of asymptomatic humans has transmitted vCJD, and 2) in a variety of experimentally TSE-infected animals, TSE agent has been detected in blood, mainly in nucleated cells and plasma (4–6,14–17). Fortunately, no human vaccine has ever been implicated as a source of iatrogenic TSE.

However, 2 animal tissue–derived vaccines have caused outbreaks of scrapie in sheep, and 2 medical products of human origin—dura mater allograft and human cadaveric pituitary hormones (no longer marketed in the United States)—have transmitted hundreds of cases of CJD; corneal grafts have transmitted a few cases as well (2,18). Since 1993, the US Food and Drug Administration has recommended against the use in the manufacture of biological products of bovine-derived materials from countries identified by the US Department of Agriculture (USDA) as having BSE or being at increased risk for BSE in native cattle (19).
The recognition of >20 BSE cases in North America since 2003 (most in Canada) has increased the need to determine whether cell substrates that might be accidentally exposed to the BSE agent are capable of acquiring and propagating the infectious agent and potentially transmitting infections to vaccine recipients (20). To address these issues, we investigated the susceptibility of cell lines used or proposed for manufacture of biologics and controls to propagate TSE agents, especially the BSE agent, under simulated worst-case conditions.

Suggested citation for this article: Piccardo P, Cervanakova L, Vasilyeva I, Yakovleva O, Bacik I, McKenzie C, et al. Candidate cell substrates, vaccine production, and transmissible spongiform encephalopathies. Emerg Infect Dis [serial on the Internet]. 2011 Dec [date cited]. http://dx.doi.org/10.3201/eid1712.110607