Summary of studies based on passive immunization against prion diseases.
Prion diseases are invariably lethal neurodegenerative diseases, associated with the structural conversion of the cellular isoform of the prion protein to its pathological, disease-associated isoform. The cellular isoform of the prion protein is highly conserved and virtually ubiquitously expressed; nevertheless, its physiological role remains unclear. Mounting evidence suggests its involvement in the regulation and function of the immune system. At the same time, the immune system is heavily involved in the pathogenesis of the diseases, playing a major role in the peripheral replication of the infectious agent and spread toward the central nervous system. On the other hand, immunotherapies are among the most promising means of intervention. This chapter deals with these fascinating and sometimes contrasting aspects of prion biology, with an emphasis on the immunization protocols developed for prophylaxis and treatment of prion diseases.
- active immunization
- passive immunization
- DNA vaccines
- mucosal vaccination
Transmissible spongiform encephalopathies (TSEs) or prion diseases are invariably lethal neurodegenerative diseases afflicting a wide variety of species, including humans . The common pathogen to all TSEs is termed prion and is believed to consist solely or primarily of the disease-associated isoform (PrPSc) of the cellular prion protein (PrPC). PrPC is a highly conserved, GPI-anchored sialoglycoprotein encoded by the single-copy
The physiological and pathological PrP isoforms display distinct conformations. The N-terminal region of PrPC is highly unstructured, as opposed to the globular C-terminal region, which contains predominantly a-helices and only a minor region encompassing two-stranded β sheet . Infrared spectroscopy and circular dichroism data indicate clear differences in the secondary structure of PrPC and PrPSc, in which equal amounts of a-helices and β sheets can be found . This conformational difference is believed to be at the basis of the biochemical differences observed between the two isoforms, namely, the partial proteinase K resistance, the reduced solubility, and the fibril-producing potential displayed by PrPSc . To date, the only known difference at the chemical level is associated with the oxidation level of the methionine residues, which was found to be elevated in PrPSc compared to PrPC .
The physiological role of the prion protein remains obscure. Its high level of conservation among species would indicate that PrPC is of crucial importance to the organism; however, PrP−/− mice are viable, developmentally and behaviorally normal, and do not display a prominent phenotype except for the complete protection against prion diseases [6, 7]. PrPC has since been implicated in a variety of cellular functions, including cell proliferation, differentiation and survival, protection against oxidative stress, and synaptic function (reviewed in [8, 9]). Further evidence suggests it may play a role in the immune system. In line with this, it has been recently reported that PrP−/− mice display lower numbers of CD4 T cells and lymphoid tissue inducer (LTi) cells as well as impaired splenic T zone structures . Moreover, immune responses have been reported during prion diseases progression, suggesting the involvement of the immune system in disease pathogenesis, and immune-based approaches have yielded some of the most promising results toward protection and/or treatment of spongiform encephalopathies. In this chapter these exciting aspects of prion biology will be discussed.
2. PrP and the immune system
2.1. PrPC expression patterns in cells of the immune system
Even though PrPC is predominantly expressed in the central and peripheral nervous system [11, 12], elevated protein expression levels have also been reported in many cells of the immune system. In long-term hematopoietic stem cells (HSCs), PrPC expression levels are raised and PrPC has been suggested as a marker for these cells . PrPC expression is retained throughout maturation either toward the myeloid  or the lymphoid lineage [15, 16]. Interestingly, along the granulocyte maturation lineage, PrPC expression is downregulated .
Among cells of the lymphoid lineage, T cells, monocytes, and natural killer (NK) cells express higher PrPC levels compared to B lymphocytes . PrPC expression levels are regulated and can vary greatly across different T-cell subtypes: CD8+ cells display higher expression levels than CD4+ cells, and between CD4+ cells, CD25+ expresses 4.5-fold higher
2.2. PrPC function in the immune system
Despite evidence that PrPC may be associated with the function of the immune system, its role remains unclear. PrP−/− mice do not display gross overt effects, at least under normal conditions. However, evidence indicates that when PrP−/− mice are subject to immunological stress their phenotype may deviate from normal. To test whether PrPC may act as a regulator of cellular immunity, the effect PrPC deficiency may have on the course of experimental autoimmune encephalomyelitis (EAE) was assessed . EAE is an inflammatory demyelinating disease of the central nervous system (CNS), triggered by the injection of brain extracts, proteins of the CNS such as the myelin basic protein and the myelin oligodendrocyte glycoprotein (MOG) or peptides from these proteins to experimental animals, usually mice and rats. EAE is widely used as an animal model for multiple sclerosis and acute disseminated encephalomyelitis but is also considered the prototype for T-cell–mediated autoimmune disease in general . It was found that PrP−/− mice displayed a more aggressive disease onset and no clinical improvement during the chronic phase of the disease. These clinical findings were in agreement with the increased cytokine gene expression in MOG-primed PrP−/− cells and indicate that PrPC could be involved in the attenuation of T-cell-dependent neuroinflammation.
Similar results were obtained when
3. Prion disease pathogenesis and the immune system
The central event in the pathogenesis of all forms of transmissible spongiform encephalopathies is the conversion of PrPC to the more thermodynamically stable PrPSc by PrPSc via a mechanism which remains at large obscure . Regrettably, the actual conversion mechanism is not the only missing piece of the prion disease pathogenesis puzzle, and not much is known on how the infectious agent enters the host or how it is transported from the periphery to the central nervous system. A series of experiments using animal models of TSEs have provided interesting data on pathogenesis.
Parenteral—usually intracranial or intraperitoneal—administration of the pathogen to hamsters or mice is among the most widely used animal TSE models. Such models are particularly useful, since most of the naturally acquired TSE cases both in humans and animals are contracted via peripheral—through the alimentary tract—exposure to the pathogen . While these models provide a wealth of information regarding pathogenesis, it later became evident that different mechanisms are involved in the pathogenesis of prion infection following the intraperitoneal or the oral route of infection , and other factors such as the pathogen strain and the host species and/or strain can also have a major impact on the mechanisms involved . For example, in a recent study in sheep with different
Prion pathogenesis can be divided into phases, some of which may take place in parallel: (i) peripheral prion exposure and uptake, (ii) peripheral pathogen replication, (iii) migration through the peripheral nervous system to the CNS, and (iv) centrifugal spread from the CNS back to the periphery [25, 27]. Despite PrPSc can be detected in various sites following peripheral exposure, especially in the lymphatic system, signs of pathology, including neurodegeneration, spongiosis, and gliosis are only found within the CNS. It is important to stress that as the means available evolve, our understanding of the phenomena taking place also improves. For instance, detection of PrPSc in the brains of some peripherally challenged hamsters as early as 4 and 9 days following challenge was recently reported .
M cells, which are epithelial cells specialized for transepithelial transport found in the follicle-associated epithelia of the small and large intestines, tonsils and adenoids , were shown capable of transcytosing the TSE infectious agent
It is not yet clear how the pathogen is transported from the entry site to the lymphoid tissue. It has been hypothesized that following pathogen uptake by M cells, the infectious agent can be transported to the M cells’ intraepithelial pocket, where it can be processed by macrophages, B- and T- lymphocytes residing within this pocket or the dendritic cells, macrophages, and lymphocytes situated immediately beneath the intraepithelial pocket . Of these cells, macrophages and dendritic cells appear the most plausible candidates for effective transport of the pathogen. In line with this assumption, PrP accumulations were detected in various types of macrophages following TSE infection [36–38]. However, the role macrophages undertake remains obscure, as
B lymphocytes were initially identified as the cells involved in replication of the TSE infectious agent , but this hypothesis was later revised, and the role of B lymphocytes in prion pathogenesis was associated with the regulated maturation of follicular dendritic cells (FDCs) . Initial experiments with splenectomized or thymectomized mice indicated the dispensable role of T lymphocytes in the replication of the agent , whereas fractionation  and irradiation  experiments indicated that replication of the pathogen depends on radioresistant cells, localized within the stromal compartment of the spleen. FDCs fulfill all these criteria, and their crucial role for replication of the pathogen was confirmed in a series of experiments, in which depletion of mature FDCs led to prolongation of the incubation period of the disease [46, 50–53]. FDCs are of stromal origin, reside in the primary B lymphocytes follicles and germinal centers of lymphoid tissues, and are non-phagocytic and non-migratory. As a result of their large surface area and longevity, FDCs are capable of trapping and retaining antigen in its native state for months to years. FDCs retain antigens in the form of immune complexes, consisting of antigen-complement components and/or antibody and trap these complexes either through complement receptors CR1 and CR2 or through FcRIIb and FcεRII antibody receptors . In agreement with the role FDCs undertake in prion pathogenesis and the involvement of complement components and receptors in antigen trapping by FDCs, it was found that the absence of complement components (C1q, C2, C3, and factor B) and cellular complement receptor can have an adverse effect on the accumulation of PrPSc in the spleen [54, 55]. However, the inability to completely inhibit disease progression via depletion of mature FDCs [46, 51], in addition to observations confirming that propagation of prion diseases is possible even in the absence of mature FDCs [41, 56–58], indicates that possibly a different cell type—most probably MOMA-1-positive macrophages —is responsible for replication of the pathogen. These diﬀerences in the cell types required for pathogenesis were attributed to the dose and agent strain .
Peripheral replication of the pathogen precedes neuroinvasion, during which the pathogen is transported within the CNS. Both the enteric and autonomic nervous systems are believed to participate in the transport of the infectious agent [36, 59, 60]. The exact mechanism governing transport of the pathogen to the CNS remains unidentified, and has been reported to be both PrPC-dependent [61, 62] and independent . Interestingly, it was reported that the transfer speed of intraperitoneally administered prions relies to the distance between FDCs and splenic nerve endings [64, 65].
The immune system is greatly implicated in the peripheral pathogenesis of prion diseases but fails to provide protection. Until recently, no response against the prion pathogen has been described, and this was linked to tolerance effects due to widespread expression of the physiological isoform of the prion protein throughout the body, which prevents the host from mounting a humoral or cellular immune response against PrPSc following infection . On the contrary, PrP−/− mice mount a robust immune response against PrP, indicating the immunogenicity of the protein. Lately it was shown that TSE infection can have adverse effects on the maturation cycle of FDCs, causing an abnormality in immune function . Given the crucial role the immune system plays in the peripheral pathogenesis of prion diseases, it could be argued that it promotes rather than protects against prion pathogenesis. In agreement with this, increased susceptibility to intraperitoneal challenge with TSE agents following repetitive immunization was recently reported .
4. Harnessing the immune system against prion diseases
Since the immune system plays an ambivalent role in prion disease pathogenesis, the question emerged whether suppressing the immune system would be the most appropriate approach . Targeting the FDCs was already proven a viable approach, providing partial protection in an animal model of prion diseases and minimizing the infectivity of the peripheral tissue of the afflicted animals [46, 50–52]. Disruption of the FDCs also appears to be the protective mechanism against TSEs following repetitive CpG administration . CpG had previously been administered as a stimulator of innate immunity and was shown effective at providing partial protection in an animal model of TSEs . In this case CpG was administered to stimulate the macrophages and enhance phagocytosis of the pathogen. Indeed, repetitive administration provided partial protection against TSEs , but as it was later shown, this protection was due to disruption of the lymphoid follicles rather than stimulation of the macrophages . Interestingly, disruption of the FDCs has also been observed following immunization of wild-type mice with recombinant murine PrP aggregates and is at least in part responsible for the observed partial protection when the immunized mice were challenged with a murine strain of TSEs .
The first indications that the immune system might prove effective against prion diseases stemmed from
From this initial series of experiments, valuable conclusions emerged, most importantly, that immunization is an efficient means of therapy rather than protection, against prion diseases. Moreover, the safety of these procedures was confirmed, since immunization against a self-antigen could always give rise to autoimmunity. Given the identical primary structure PrPC and PrPSc share, adverse reactions stemming from the reaction of the anti-PrP antibodies with PrPC could be expected. Autoimmunity was not induced by these immunization approaches, and furthermore the “dispensable” role of the prion protein for the appearance of a physiological phenotype was already known from studies on PrP-/- animals , as well as from transgenic animals with conditional depletion of the prion protein  and provided an extra layer of security. However, other findings raised some concerns over the safety of administration of anti-PrP antibodies, since it was found that intracerebral administration of anti-PrP monoclonal antibodies can give rise to cross-linking of PrP molecules on adjacent neurons and eventually cell death, triggered possibly by the initiation of death signaling . These effects are clearly not associated with autoimmunity, but rather with impaired cell signaling.
4.1. Passive immunization approaches
The first indications that passive immunization could prove useful at protecting against prion diseases emerged from studies in which mice genetically modified to produce an anti-PrP monoclonal antibody (6H4μ) were fully protected against prion diseases . In a more classical approach, monoclonal anti-PrP antibodies (ICSM18 and ICSM35) were administered intraperitoneally to wild-type mice briefly after intraperitoneal inoculation with the pathogen or when the first clinical signs appeared. When the antibodies were administered after the inoculation, animals receiving the antibodies survived approximately 300 days more than control mice, and the accumulation of infectivity in the peripheral tissue was markedly reduced . Intraperitoneal administration of a different antibody (6D11) immediately after intraperitoneal administration of the pathogen also proved its protective efficacy, since mice receiving the antibody survived longer by approximately 36.9% compared to control mice. In a recent study, a pharmacokinetic and pharmacodynamic analysis following intraperitoneal administration of various anti-PrP antibodies was carried out. The ability of an antibody to form long-lasting complexes with PrPC was found to positively correlate with its efficacy in delaying peripheral accumulation of PrPSc and, in agreement with this, intraperitoneal administration of the monoclonal antibody BAR216 led to a statistically significant prolongation of survival of the mice .
The therapeutic efficacy of intracerebral administration of anti-PrP monoclonal antibodies was evaluated in two recent studies. In the first one, monoclonal antibody 4H11 (F(ab′)2 and IgG) was intraventricularly administered to transgenic mice overexpressing PrP using osmotic pumps from d85 to d100 following intraperitoneal challenge with a mouse-adapted bovine spongiform encephalopathy (BSE) strain. The mice were not protected by this regimen, and they succumbed to disease concomitantly with the control mice. Furthermore, mice treated with the antibodies developed neuronal cell death, associated with administration of the antibodies. In addition to previously reported results , linking cell death to PrP cross-linking events, in this study, emerged that PrP cross-linking is not the only mechanism mediating cell death; “coating” the whole cell surface PrP with antibodies or antibodies fragments could induce other toxic signals . In the second study, intraventricular administration of antibodies 106, 110, 31C6, and 44B1 to wild-type mice was not linked with neuronal cell death; however, only a minor prolongation of survival and in one of the two tested animal models was achieved following administration of the monoclonal antibodies . Differences in the epitopes recognized by the antibodies used in these two studies as well as the use of PrP overexpressing versus wild-type mice could account for the different results obtained regarding neuronal cell death. Of note, neuronal cell death has been challenged in another, more recent study, and it would be safe to assume that toxic effects are associated with the epitope and the dosage of the antibodies used .
A completely different passive immunization approach was used in two other studies; based on the discovery of the non-integrin 37/67 kDa lamin receptor (LRP/LR) as an interaction partner for both isoforms of PrP [84–86], polyclonal anti-LRP/LR  or single-chain Fv anti-LRP/LR antibodies  were intraperitoneally administered to wild-type mice as protective means in a mouse model of prion diseases. On both occasions, peripheral PrPSc accumulation was reduced; however, partial protection was only achieved with the polyclonal antibodies. This difference in the efficacy was attributed to differences in the pharmacokinetics and dosage regimen; polyclonal antibodies have a half-life of approximately 14 days in the blood, whereas the single-chain antibodies have a half-life of only 12 h. Moreover, the polyclonal antibodies were administered for 12 weeks, starting 1 week before administration of the pathogen, whereas the single-chain antibodies for 8 weeks. Passive immunization approaches are summarized in Table 1 .
|Antibody name||Antibody type and target||Epitope||Administration protocol||Reference|
|ICSM18, ICSM35||Monoclonal, PrP||ICSM18: 143–153aa|
|Intraperitoneal administration twice weekly starting 7 or 30 days after administration of the pathogen or at onset of the clinical symptoms||NP||Prolonged survival interval in a mouse model of prion disease when the antibodies were administered prior to the appearance of clinical symptoms|||
|6D11||Monoclonal, PrP||97–110aa||One intravenous administration immediately after administration of the pathogen followed by consecutive intraperitoneal administrations (twice per week for 4 or 8 weeks)||Prevention of infection and clearance of infection in already prion-infected cell lines||Prolongation of incubation period in a mouse model of prion disease|||
|BAR236||Monoclonal, PrP||Linear epitope unidentified||Intraperitoneal (3 weekly administrations, starting 1 week after administration of the pathogen)||NP||Prolongation of survival interval in a mouse model of prion disease|||
|4H11||Monoclonal or F(ab′)2 fragments, PrP||Epitope within octarepeat region (59–89aa)||Intraventricular (osmotic pump delivering antibody for 16 days starting 85 days after administration of the pathogen)||Inhibition of PrPSc propagation in an already prion-infected cell line. Recognition of PrP on the cell surface by FACS||Intraventricular administration of the antibody did not prolong survival interval in a mouse model of prion disease|||
|106, 110, 31C6, 44B1||Monoclonal, anti-PrP||106: 88–90aa|
44B1: discontinuous epitope within aa 155–231 aa
|Intraventricular (osmotic pump delivering antibody for 14 days starting 60, 90, or 120 days after administration of the pathogen)||NP||Small (8%) prolongation of survival interval in a mouse model of prion diseases, even when administration of antibodies commences after appearance of first symptoms (120 days after administration of the pathogen)|||
|pAb W3||Polyclonal anti-LRP/LR||Undefined||Intraperitoneal (12 weekly administrations starting 1 week before administration of the pathogen)||NP||Prolongation of survival interval, but not of incubation period in a mouse model of prion disease|||
|S18||scFV, LRP||272–280aa||Intraperitoneal (8 weekly administrations starting 1 day before administration of the pathogen)||S18 prevents interaction of the recombinant human PrP with recombinant human LRP||Reduction of splenic PrPSc, but no prolongation of survival interval in a mouse model of prion disease|||
|W226||Monoclonal, scFV||Undefined||Intraperitoneal administration twice weekly starting 2 or 28 days after administration of the pathogen or at onset of the clinical symptoms||Clearance of PrPSc in ScN2a cells||Minor delay of incubation time in immunized versus control mice|||
|EB8, DC2, DE10, EF2||Monoclonal||EB8: 26–34aa; DC2: 35–46aa;|
DE10: 44–52aa and EF2: 47–52aa
|NP||Clearance of PrPSc in ScGT2 cells||NP|||
4.2. Active immunization approaches
Although passive immunization does protect against prion diseases, it provides a narrow window for intervention, i.e., antibodies must be administered shortly after exposure to the pathogen. In this regard, active immunization against the prion protein, which provides protection against the diseases similarly to a conventional vaccine, could prove a much more useful approach. Nevertheless, the prion protein-associated tolerance effects which prevent the immune system from mounting an immune response against the prion protein hinder development of such approaches .
Despite the tolerance effects, initiation of a humoral immune response against the prion protein was achieved, albeit with mediocre results in terms of protection against the disease. In the first reports, wild-type mice were immunized with recombinant murine prion protein mixed with complete Freund’s adjuvant (CFA) and challenged with a mouse-adapted scrapie strain either concomitantly with the immunization (rescue treatment) or following its completion (prophylactic treatment). Although the mice developed antibodies against the prion protein, only mice of the prophylactic treatment group were partially protected against the pathogen; mice of this group succumbed to disease with a delay of approximately 16d compared to control mice .
4.2.1. Peptide-based active immunization
Numerous strategies were implemented to overcome the tolerance effects and promote generation of anti-prion antibodies. The most obvious approach was to use prion peptides properly modified to enhance the antigenicity of the protein (summarized in Table 2 ). Following this rationale, wild-type animals were immunized with prion protein peptides [90–93], PrP dimers [94–96], or PrP aggregates . In addition to homologous prion protein immunization , which provided proof of principle that active immunization can have a protective role against prion diseases, immunization with heterologous prion peptides also provided rather encouraging results . In an attempt to enhance the immunogenicity of the prion peptides, various adjuvants, including Freund’s adjuvant, Montanide IMS-1313, TiterMax, CpG, anti-OX40 antibodies—antibodies against the signaling molecule CD134, which recently has been shown to break T cell tolerance—and keyhole limpet hemocyanin, were used [95, 98], as well as different vaccine formulations, including encapsulation of the CpG-antigen complex in polylactide-coglycolide microspheres . Interestingly, an early report indicates that immunization with complete Freund’s adjuvant alone can provide partial protection in a mouse model of prion diseases through an unidentified mechanism . Based on the extremely strong adjuvant effect exerted by heat-shock proteins, PrP molecules chemically cross-linked  or fused  to recombinant bacterial heat-shock proteins were also used to immunize wild-type mice and lead to the production of antibodies that recognized recombinant PrP.
|Antigen||Animals immunized||Humoral response||T-cell responses||Reference|
|Various murine PrP peptides||Wild-type mice||+||NP||NP||Reduction of proteinase K-resistant prion protein in a scrapie-infected tumor transplant|||
|Recombinant murine PrP chemically cross-linked to bacterial heat-shock proteins||Wild-type mice||+||NP||NP||NP|||
|Recombinant murine PrP||Wild-type mice||+||NP||NP||Prolongation of survival interval in a mouse model of prion disease|||
|Recombinant murine PrP dimer||Wild-type mice, rabbits||+||NP||Polyclonal sera produced reduced PrPSc synthesis in prion-infected cell lines||NP|||
|Recombinant murine prion peptide 105–125 linked to keyhole limpet hemocyanin and recombinant murine prion 90–230||Wild-type mice||+||NP||NP||Prolongation of survival interval in a mouse model of prion disease|||
|Mouse prion peptides 31–50 and 211–230||Wild-type mice||NP||NP||NP||Prolongation of survival interval in a mouse model of prion disease, even when only the adjuvant Complete Freund’s Adjuvant (CFA) is administered|||
|Various murine prion peptides and adjuvants||Wild-type mice||+||ND||FACS to detect binding of the produced antibodies on native PrP||Statistically insignificant prolongation of survival time in a mouse model of prion disease|||
|Murine prion peptides 39–67, 98–127, 143–172, and 158–187 with CFA or CpG||Wild-type mice||+||+||NP||NP|||
|Hamster prion peptides 105–128, 119–146, and 142–179||Wild-type hamsters||+||NP||NP||Prolongation of survival interval in a hamster model of prion diseases|||
|Recombinant murine, ovine, and bovine prion protein||Wild-type mice||Detected following immunization with ovine and bovine recombinant PrP||NP||NP||Prolongation of survival interval in a mouse model of prion diseases following immunization with the bovine-recombinant protein|||
|Recombinant murine PrP dimer and CpG encapsulated in polylactide-coglycolide microspheres||Wild-type mice||+||+||NP||NP|||
|Murine scrapie-associated fibrils and CpG||Transgenic and wild-type mice||+||NP||NP||Prolongation of the survival interval of the wild-type mice in a mouse model of prion disease when CpG was used|||
|Murine scrapie-associated fibrils immobilized on Dynabeads||Wild-type mice||+||NP||NP||Prolongation of survival interval in a mouse model of prion diseases with the bovine-recombinant protein|||
|Cervid prion peptide sequences 168–182 and 145–164||Deer||+||NP||NP||Delay of incubation time in immunized versus control mice|||
|Prion disease-derived brain material||Camelid||+||NP||Permanent abrogation of prion replication in a prion-permissive cell line||NP|||
|rPrP aggregates, solubilized rPrP, DnaK-fused PrP||Mouse||+||NP||FACS to detect binding of the produced antibodies on native PrP||Statistically significant prolongation of survival time in a mouse model of prion disease|||
Despite the widely accepted notion that PrPSc is not immunogenic and that the immune system does not provide protection against PrPSc in wild-type animals, when highly purified proteinase K-resistant PrPSc, originating from murine brains afflicted with an animal model of prion diseases was coadministered with CpG  or administered immobilized on Dynabeads coated with antibodies against PrP  a humoral immune response, which providing partial protection in animal model of prion diseases was elicited.
Although the protective role of the aforementioned, peptide-based approaches was not investigated on all occasions, it became evident that using various approaches the self-tolerance effects can be overcome and immune reactions against the prion protein can be obtained. However, it appears that protection against TSEs is restricted to antibodies capable of recognizing the native cell-surface PrPC . This requirement was met by antibodies known to provide protection against TSEs, e.g., ICSM18  and 6H4 , whereas other antibodies capable of recognizing recombinant PrP but unable to provide protection against TSEs also failed to recognize native PrPC [72, 95].
4.2.2. DNA vaccines
In addition to peptide-based vaccines, DNA vaccines were also used to promote immune responses against the prion protein. In this case, nucleic acid encoding for the prion protein is administered to animals, wherein the nucleic acid is translated to the corresponding protein and an immune response is initiated. The first attempt at raising anti-PrP antibodies using DNA vaccines was only successful in PrP−/− mice, whereas the same approach failed to give rise to anti-PrP antibodies in wild-type mice . Induction of anti-PrP antibodies using DNA vaccines in wild-type mice was triggered when the mice were immunized with a DNA construct coding for the murine prion protein fused to the lysosomal targeting signal from lysosomal integral membrane protein type II (LIMPII). Immunization with this construct leads to a remarkable delay on the onset of disease symptoms, which was not followed by a similar prolongation of survival interval. This discrepancy in the obtained results was attributed to immunopathology mediated by PrP-specific antibodies induced by the DNA vaccine used and constitutes the first report of adverse effects following active prion immunization .
In a different approach, DNA vaccines were used to prime wild-type mice, followed by peptide immunizations to further boost immune responses. Although this approach was successful when PrP-/- mice were immunized, very low antibody titers and only marginal protection were achieved when tested on wild-type mice . In a recent report, wild-type mice were immunized with cDNA coding for human PrPC fused to a T-cell stimulatory peptide. These mice developed a strong humoral immune response against the native protein, and although a bioassay was not carried out, the produced antibodies were capable of recognizing the native conformation of murine PrPC, which—as already mentioned—constitutes a strong indicator of protective efficiency against prion diseases . Studies based on DNA vaccines are summarized in Table 3 .
|Vaccine||Immunized animals||Humoral response||T-cell responses||Reference|
|DNA vaccine encoding either murine PrP or murine PrP fused to ubiquitin or to a lysosomal targeting signal||Wild-type mice||+||+||NP||Prolongation of asymptomatic period and accumulation of disease associated PrP, but not of survival interval. Death of the immunized mice was attributed to neurodegeneration associated with production of anti-PrP antbodies|||
|DNA vaccine encoding murine PrP linked to helper T-cell epitopes|
Combination of DNA and peptide immunization
|PrP−/− and wild-type mice||Achieved in PrP−/− mice, very low titer in wild-type mice||Detected in PrP−/− mice but not wild-type mice||FACS to detect binding of the produced antibodies on native PrP positive with PrP−/− mice sera, negative with wild-type mice sera. PrP−/− mice sera reduced PrPSc levels in prion-infected cell lines||Not effective|||
|DNA vaccine encoding human PrP fused or not to a tetanus toxin stimulatory T-cell epitope and PrP protein boost||Wild-type mice||+||NP||FACS to detect binding of the produced antibodies on native PrP||NP|||
|DNA vaccine encoding human PrP fused to ubiquitin, lysosomal integral membrane protein type II lysosome‐targeting signal or an ER-targeting signal in conjunction with PrP vaccination||Wild-type mice||+||+||NP||NP|||
4.2.3. Immunization with PrP-displaying viral constructs
A different approach to overcome the tolerance effects and stimulate the production of anti-PrP antibodies in wild-type mice is the expression of the prion protein on the surface of viral particles (summarized in Table 4 ). Virus-like particles (VLPs) are much better B lymphocytes immunogens than monovalent proteins and would be expected to trigger a stronger humoral immune response by passing tolerance.
|Vaccine||Immunized animals||Humoral response||T-cell responses||Reference|
|Murine PrP or C-terminal murine PrP expressed on recombinant retroviral virus-like particles||PrP−/− and wild-type mice||+||NP||FACS to detect binding of the produced antibodies on native PrP||NP|||
|Murine/rat prion 9/mer inserted into the L1 major capsid protein of bovine papillomavirus type 1||Wild-type rabbits and rats||+||NP||FACS to detect binding of the produced antibodies on native PrP, immunoprecipitation|
Rabbit immune sera inhibited de novo synthesis of PrPSc in prion-infected cells
|Priming with adenovirus 5 expressing the human PrP gene followed by boosting with the human PrP plasmid||PrP−/− and wild-type mice||+||+||FACS to detect binding of the produced antibodies on native PrP||Marginal prolongation of survival interval of the immunized mice|||
In a first attempt, retroviral particles displaying the C-terminal portion of murine PrP were used to immunize wild-type mice. These mice developed anti-PrP antibodies, capable of recognizing the native form of PrPC, thus displaying strong therapeutic potential . A similar approach was used to insert the 9-amino-acid-, prion-pathogenesis associated-peptide pertaining to the murine/rat prion protein into the L1 major capsid protein of bovine papillomavirus type 1. These VLPs were used to immunize both wild-type rats and rabbits. The anti-sera collected from both immunized species recognized native PrPSc, and importantly immune serum from the immunized rabbit prevented synthesis of PrPSc in scrapie-infected cell lines . In a more recent approach, dendritic cells transduced with adenoviruses encoding the human prion protein were used to immunize wild-type mice. These mice developed antibodies against the murine prion protein as well, which provided partial protection against TSEs, as shown by the reduction in splenic PrPSc accumulation and prolongation of survival interval in a murine model of TSEs .
4.2.4. Mucosal immunization
To date, the only active immunization strategy providing complete protection against prion diseases is mucosal immunization. To trigger mucosal immunization, either transgenic, live-attenuated
To induce mucosal immunization, a live-attenuated
|Vaccine||Immunized animals||Humoral response||T-cell responses||Reference|
|Orally administered ||Wild-type mice||+||NP||NP||Significant prolongation of survival interval in a mouse model of prion disease||[110, 112]|
|Intranasally, intragastrically, or intraperitoneally administered murine PrP90–231 and cholera toxin||Wild-type mice||+||NP||NP||Marginal prolongation of survival interval in a mouse model of prion disease following intranasal administration|||
|Orally administered ||White-tailed deers||+||NP||NP||Significant prolongation of survival interval in an elk model of prion disease. One immunized animal remained asymptomatic|||
Although mucosal immunization is only effective following oral exposure, it is important to remember that the gut is the major route of entry for prion diseases such as CWD in white-tailed deer, BSE in cattle, and variant Creutzfeldt-Jakob disease and kuru in humans. Furthermore, mucosal vaccination can be properly designed to induce a primarily humoral immune response and is unlikely to produce a significant immune response within the brain, thus minimizing the risk of appearance of adverse reactions .
5. Future perspectives
Despite fervent research and some very encouraging results, many facets of the involvement of the immune system in prion pathogenesis remain obscure, and a powerful immunoprotective tool has yet to emerge. Passive immunization with anti-prion antibodies and mucosal immunization were the only two approaches to provide satisfactory results but have a series of limitations associated with the narrow window of intervention and the route of infection. However, immune-based therapeutics both in their more classical immunization-based form or more modern, immunomodulatory form  hold great promise for prion diseases and other protein-misfolding diseases.
Prusiner, S.B., Novel proteinaceous infectious particles cause scrapie. Science, 1982. 216(4542): pp. 136–144.
Riek, R., et al., NMR characterization of the full- length recombinant murine prion protein, mPrP( 23- 231). FEBS Lett, 1997. 413(2): pp. 282–288.
Caughey, B.W., et al., Secondary structure analysis of the scrapie- associated protein PrP 27- 30 in water by infrared spectroscopy. Biochemistry, 1991. 30(31): pp. 7672–7680.
Prusiner, S.B., Prions. Proc Natl Acad Sci U S A, 1998. 95(23): pp. 13363–13383.
Canello, T., et al., Methionine sulfoxides on PrPSc: a prion- specific covalent signature. Biochemistry, 2008. 47(34): pp. 8866–8873.
Bueler, H., et al., Normal development and behaviour of mice lacking the neuronal cell- surface PrP protein. Nature, 1992. 356(6370): pp. 577–582.
Manson, J.C., et al., 129/ Ola mice carrying a null mutation in PrP that abolishes mRNA production are developmentally normal. Mol Neurobiol, 1994. 8(2–3): pp. 121–127.
Aguzzi, A., Baumann, F., and Bremer, J., The prion’ s elusive reason for being. Annu Rev Neurosci, 2008. 31: pp. 439–477.
Linden, R., et al., Physiology of the prion protein. Physiol Rev, 2008. 88(2): pp. 673–728.
Kim, S., et al., Prion protein- deficient mice exhibit decreased CD4 T and LTi cell numbers and impaired spleen structure. Immunobiology, 2016. 221(1): pp. 94–102.
Chesebro, B., et al., Identification of scrapie prion protein- specific mRNA in scrapie- infected and uninfected brain. Nature, 1985. 315(6017): pp. 331–333.
Oesch, B., et al., A cellular gene encodes scrapie PrP 27- 30 protein. Cell, 1985. 40(4): pp. 735–746.
Zhang, C.C., et al., Prion protein is expressed on long- term repopulating hematopoietic stem cells and is important for their self- renewal. Proc Natl Acad Sci U S A, 2006. 103(7): pp. 2184–2189.
Burthem, J., et al., The normal cellular prion protein is strongly expressed by myeloid dendritic cells. Blood, 2001. 98(13): pp. 3733–3738.
Cashman, N.R., et al., Cellular isoform of the scrapie agent protein participates in lymphocyte activation. Cell, 1990. 61(1): pp. 185–192.
Li, R., et al., The expression and potential function of cellular prion protein in human lymphocytes. Cell Immunol, 2001. 207(1): pp. 49–58.
Dodelet, V.C. and Cashman, N.R., Prion protein expression in human leukocyte differentiation. Blood, 1998. 91(5): pp. 1556–1561.
Durig, J., et al., Differential constitutive and activation- dependent expression of prion protein in human peripheral blood leucocytes. Br J Haematol, 2000. 108(3): pp. 488–495.
Isaacs, J.D., Jackson, G.S., and Altmann, D.M., The role of the cellular prion protein in the immune system. Clin Exp Immunol, 2006. 146(1): pp. 1–8.
Tsutsui, S., et al., Absence of the cellular prion protein exacerbates and prolongs neuroinflammation in experimental autoimmune encephalomyelitis. Am J Pathol, 2008. 173(4): pp. 1029–1041.
Miller, S.D. and Karpus, W.J., Experimental autoimmune encephalomyelitis in the mouse. Curr Protoc Immunol, 2007. Chapter 15: p. Unit 15 1.
Hu, W., et al., Pharmacological prion protein silencing accelerates central nervous system autoimmune disease via T cell receptor signalling. Brain, 2010. 133(Pt 2): pp. 375–388.
Bakkebo, M.K., et al., The cellular prion protein: a player in immunological quiescence. Front Immunol, 2015. 6: p. 450.
Riesner, D., Biochemistry and structure of PrP( C) and PrP( Sc). Br Med Bull, 2003. 66: pp. 21–33.
Beekes, M. and McBride, P.A., The spread of prions through the body in naturally acquired transmissible spongiform encephalopathies. FEBS J, 2007. 274(3): pp. 588–605.
Prinz, M., et al., Oral prion infection requires normal numbers of Peyer’ s patches but not of enteric lymphocytes. Am J Pathol, 2003. 162(4): pp. 1103–1111.
Aguzzi, A. and M. Heikenwalder, Pathogenesis of prion diseases: current status and future outlook. Nat Rev Microbiol, 2006. 4(10): pp. 765–775.
Greenlee, J.J., et al., Lack of prion accumulation in lymphoid tissues of PRNP ARQ/ ARR sheep intracranially inoculated with the agent of scrapie. PLoS One, 2014. 9(9): p. e108029.
Friedman-Levi, Y., et al., Genetic prion disease: no role for the immune system in disease pathogenesis?. Hum Mol Genet, 2014. 23(15): pp. 4134–4141.
Chen, B., Soto, C., and Morales, R., Peripherally administrated prions reach the brain at sub- infectious quantities in experimental hamsters. FEBS Lett, 2014. 588(5): pp. 795–800.
Kraehenbuhl, J.P. and Neutra, M.R., Epithelial M cells: differentiation and function. Annu Rev Cell Dev Biol, 2000. 16: pp. 301–332.
Heppner, F.L., et al., Transepithelial prion transport by M cells. Nat Med, 2001. 7(9): pp. 976–977.
Mishra, R.S., et al., Protease- resistant human prion protein and ferritin are cotransported across Caco- 2 epithelial cells: implications for species barrier in prion uptake from the intestine. J Neurosci, 2004. 24(50): pp. 11280–11290.
Donaldson, D.S., Else, K.J., and Mabbott, N.A., The gut- associated lymphoid tissues in the small intestine, not the large intestine, play a major role in oral prion disease pathogenesis. J Virol, 2015. 89(18): pp. 9532–9547.
Mabbott, N.A. and MacPherson, G.G., Prions and their lethal journey to the brain. Nat Rev Microbiol, 2006. 4(3): pp. 201–211.
Beekes, M. and McBride, P.A., Early accumulation of pathological PrP in the enteric nervous system and gut- associated lymphoid tissue of hamsters orally infected with scrapie. Neurosci Lett, 2000. 278(3): pp. 181–184.
Herrmann, L.M., et al., CD21- positive follicular dendritic cells: a possible source of PrPSc in lymph node macrophages of scrapie- infected sheep. Am J Pathol, 2003. 162(4): pp. 1075–1081.
Jeffrey, M., et al., Sites of prion protein accumulation in scrapie- infected mouse spleen revealed by immuno- electron microscopy. J Pathol, 2000. 191(3): pp. 323–332.
Carp, R.I. and Callahan, S.M., In vitro interaction of scrapie agent and mouse peritoneal macrophages. Intervirology, 1981. 16(1): pp. 8–13.
Sassa, Y., Inoshima, Y., and Ishiguro, N., Bovine macrophage degradation of scrapie and BSE PrP(Sc). Vet Immunol Immunopathol, 2009. 133(1): pp. 33–39.
Prinz, M., et al., Lymph nodal prion replication and neuroinvasion in mice devoid of follicular dendritic cells. Proc Natl Acad Sci U S A, 2002. 99(2): pp. 919–924.
Takahashi, K., Inoshima, Y., and Ishiguro, N., Role of cell death in the propagation of PrP( Sc) in immune cells. Arch Virol, 2015. 160(3): pp. 693–699.
Huang, F.P., et al., Migrating intestinal dendritic cells transport PrP( Sc) from the gut. J Gen Virol, 2002. 83(Pt 1): pp. 267–271.
Mohan, J., Hopkins, J., and Mabbott, N.A., Skin- derived dendritic cells acquire and degrade the scrapie agent following in vitro exposure. Immunology, 2005. 116(1): pp. 122–133.
Klein, M.A., et al., A crucial role for B cells in neuroinvasive scrapie. Nature, 1997. 390(6661): pp. 687–690.
Montrasio, F., et al., Impaired prion replication in spleens of mice lacking functional follicular dendritic cells. Science, 2000. 288(5469): pp. 1257–1259.
Fraser, H. and Dickinson, A.G., Studies of the lymphoreticular system in the pathogenesis of scrapie: the role of spleen and thymus. J Comp Pathol, 1978. 88(4): pp. 563–573.
Clarke, M.C. and Kimberlin, R.H., Pathogenesis of mouse scrapie: distribution of agent in the pulp and stroma of infected spleens. Vet Microbiol, 1984. 9(3): pp. 215–225.
Fraser, H. and Farquhar, C.F., Ionising radiation has no influence on scrapie incubation period in mice. Vet Microbiol, 1987. 13(3): pp. 211–223.
Mabbott, N.A., et al., Temporary inactivation of follicular dendritic cells delays neuroinvasion of scrapie. Nat Med, 2000. 6(7): pp. 719–720.
Mabbott, N.A., et al., Follicular dendritic cell dedifferentiation by treatment with an inhibitor of the lymphotoxin pathway dramatically reduces scrapie susceptibility. J Virol, 2003. 77(12): pp. 6845–6854.
Mohan, J., Bruce, M.E., and Mabbott, N.A., Follicular dendritic cell dedifferentiation reduces scrapie susceptibility following inoculation via the skin. Immunology, 2005. 114(2): pp. 225–234.
Mabbott, N.A., et al., Temporary blockade of the tumor necrosis factor receptor signaling pathway impedes the spread of scrapie to the brain. J Virol, 2002. 76(10): pp. 5131–5139.
Klein, M.A., et al., Complement facilitates early prion pathogenesis. Nat Med, 2001. 7(4): pp. 488–492.
Mabbott, N.A., et al., Temporary depletion of complement component C3 or genetic deficiency of C1q significantly delays onset of scrapie. Nat Med, 2001. 7(4): pp. 485–487.
Oldstone, M.B., et al., Lymphotoxin- alpha- and lymphotoxin- beta- deficient mice differ in susceptibility to scrapie: evidence against dendritic cell involvement in neuroinvasion. J Virol, 2002. 76(9): pp. 4357–4363.
Shlomchik, M.J., et al., Neuroinvasion by a Creutzfeldt- Jakob disease agent in the absence of B cells and follicular dendritic cells. Proc Natl Acad Sci U S A, 2001. 98(16): pp. 9289–9294.
Manuelidis, L., et al., Follicular dendritic cells and dissemination of Creutzfeldt- Jakob disease. J Virol, 2000. 74(18): pp. 8614–8622.
Baldauf, E., Beekes, M., and Diringer, H., Evidence for an alternative direct route of access for the scrapie agent to the brain bypassing the spinal cord. J Gen Virol, 1997. 78(Pt 5): pp. 1187–1197.
McBride, P.A., et al., Early spread of scrapie from the gastrointestinal tract to the central nervous system involves autonomic fibers of the splanchnic and vagus nerves. J Virol, 2001. 75(19): pp. 9320–9327.
Race, R., Oldstone, M., and Chesebro, B., Entry versus blockade of brain infection following oral or intraperitoneal scrapie administration: role of prion protein expression in peripheral nerves and spleen. J Virol, 2000. 74(2): pp. 828–833.
Glatzel, M. and Aguzzi, A., PrP( C) expression in the peripheral nervous system is a determinant of prion neuroinvasion. J Gen Virol, 2000. 81(Pt 11): pp. 2813–2821.
Kunzi, V., et al., Unhampered prion neuroinvasion despite impaired fast axonal transport in transgenic mice overexpressing four- repeat tau. J Neurosci, 2002. 22(17): pp. 7471–7477.
Prinz, M., et al., Positioning of follicular dendritic cells within the spleen controls prion neuroinvasion. Nature, 2003. 425(6961): pp. 957–962.
von Poser-Klein, C., et al., Alteration of B- cell subsets enhances neuroinvasion in mouse scrapie infection. J Virol, 2008. 82(7): pp. 3791–3795.
Mabbott, N.A., Prospects for safe and effective vaccines against prion diseases. Expert Rev Vaccines, 2015. 14(1): pp. 1–4.
McGovern, G., Mabbott, N., and Jeffrey, M., Scrapie affects the maturation cycle and immune complex trapping by follicular dendritic cells in mice. PLoS One, 2009. 4(12): p. e8186.
Bremer, J., et al., Repetitive immunization enhances the susceptibility of mice to peripherally administered prions. PLoS One, 2009. 4(9): p. e7160.
Aguzzi, A. and Sigurdson, C.J., Antiprion immunotherapy: to suppress or to stimulate?. Nat Rev Immunol, 2004. 4(9): pp. 725–736.
Heikenwalder, M., et al., Lymphoid follicle destruction and immunosuppression after repeated CpG oligodeoxynucleotide administration. Nat Med, 2004. 10(2): pp. 187–192.
Sethi, S., et al., Postexposure prophylaxis against prion disease with a stimulator of innate immunity. Lancet, 2002. 360(9328): pp. 229–230.
Xanthopoulos, K., et al., Immunization with recombinant prion protein leads to partial protection in a murine model of TSEs through a novel mechanism. PLoS One, 2013. 8(3): p. e59143.
Enari, M., Flechsig, E., and Weissmann, C., Scrapie prion protein accumulation by scrapie- infected neuroblastoma cells abrogated by exposure to a prion protein antibody. Proc Natl Acad Sci U S A, 2001. 98(16): pp. 9295–9299.
Peretz, D., et al., Antibodies inhibit prion propagation and clear cell cultures of prion infectivity. Nature, 2001. 412(6848): pp. 739–743.
Perrier, V., et al., Anti- PrP antibodies block PrPSc replication in prion- infected cell cultures by accelerating PrPC degradation. J Neurochem, 2004. 89(2): pp. 454–463.
Heppner, F.L., et al., Prevention of scrapie pathogenesis by transgenic expression of anti- prion protein antibodies. Science, 2001. 294(5540): pp. 178–182.
White, A.R., et al., Monoclonal antibodies inhibit prion replication and delay the development of prion disease. Nature, 2003. 422(6927): pp. 80–83.
Mallucci, G., et al., Depleting neuronal PrP in prion infection prevents disease and reverses spongiosis. Science, 2003. 302(5646): pp. 871–874.
Solforosi, L., et al., Cross- linking cellular prion protein triggers neuronal apoptosis in vivo. Science, 2004. 303(5663): pp. 1514–1516.
Feraudet-Tarisse, C., et al., Immunotherapeutic eﬀect of anti-PrP monoclonal antibodies in TSE mouse models: pharmacokinetic and pharmacodynamic analysis. J Gen Virol, 2010. 91(Pt 6): pp. 1635–1645
Lefebvre-Roque, M., et al., Toxic effects of intracerebral PrP antibody administration during the course of BSE infection in mice. Prion, 2007. 1(3): pp. 198–206.
Song, C.H., et al., Effect of intraventricular infusion of anti- prion protein monoclonal antibodies on disease progression in prion- infected mice. J Gen Virol, 2008. 89(Pt 6): pp. 1533–1544.
Klohn, P.C., et al., PrP antibodies do not trigger mouse hippocampal neuron apoptosis. Science, 2012. 335(6064): p. 52.
Gauczynski, S., et al., The 37- kDa/ 67- kDa laminin receptor acts as the cell- surface receptor for the cellular prion protein. EMBO J, 2001. 20(21): pp. 5863–5875.
Morel, E., et al., Bovine prion is endocytosed by human enterocytes via the 37 kDa/ 67 kDa laminin receptor. Am J Pathol, 2005. 167(4): pp. 1033–1042.
Gauczynski, S., et al., The 37- kDa/ 67- kDa laminin receptor acts as a receptor for infectious prions and is inhibited by polysulfated glycanes. J Infect Dis, 2006. 194(5): pp. 702–709.
Zuber, C., et al., Anti- LRP/ LR antibody W3 hampers peripheral PrPSc propagation in scrapie infected mice. Prion, 2007. 1(3): pp. 207–212.
Zuber, C., et al., Single chain Fv antibodies directed against the 37 kDa/ 67 kDa laminin receptor as therapeutic tools in prion diseases. Mol Immunol, 2008. 45(1): pp. 144–151.
Sigurdsson, E.M., et al., Immunization delays the onset of prion disease in mice. Am J Pathol, 2002. 161(1): pp. 13–17.
Arbel, M., Lavie, V., and Solomon, B., Generation of antibodies against prion protein in wild- type mice via helix 1 peptide immunization. J Neuroimmunol, 2003. 144(1–2): pp. 38–45.
Magri, G., et al., Decrease in pathology and progression of scrapie after immunisation with synthetic prion protein peptides in hamsters. Vaccine, 2005. 23(22): pp. 2862–2868.
Schwarz, A., et al., Immunisation with a synthetic prion protein- derived peptide prolongs survival times of mice orally exposed to the scrapie agent. Neurosci Lett, 2003. 350(3): pp. 187–189.
Souan, L., et al., Modulation of proteinase- K resistant prion protein by prion peptide immunization. Eur J Immunol, 2001. 31(8): pp. 2338–2346.
Gilch, S., et al., Polyclonal anti- PrP auto- antibodies induced with dimeric PrP interfere efficiently with PrPSc propagation in prion- infected cells. J Biol Chem, 2003. 278(20): pp. 18524–18531.
Polymenidou, M., et al., Humoral immune response to native eukaryotic prion protein correlates with anti- prion protection. Proc Natl Acad Sci U S A, 2004. 101Suppl 2: pp. 14670–14676.
Kaiser-Schulz, G., et al., Polylactide- coglycolide microspheres co- encapsulating recombinant tandem prion protein with CpG- oligonucleotide break self- tolerance to prion protein in wild- type mice and induce CD4 and CD8 T cell responses. J Immunol, 2007. 179(5): pp. 2797–2807.
Ishibashi, D., et al., Immunization with recombinant bovine but not mouse prion protein delays the onset of disease in mice inoculated with a mouse- adapted prion. Vaccine, 2007. 25(6): pp. 985–992.
Rosset, M.B., et al., Breaking immune tolerance to the prion protein using prion protein peptides plus oligodeoxynucleotide- CpG in mice. J Immunol, 2004. 172(9): pp. 5168–5174.
Tal, Y., et al., Complete Freund’ s adjuvant immunization prolongs survival in experimental prion disease in mice. J Neurosci Res, 2003. 71(2): pp. 286–290.
Koller, M.F., Grau, T., and Christen, P., Induction of antibodies against murine full- length prion protein in wild- type mice. J Neuroimmunol, 2002. 132(1–2): pp. 113–116.
Spinner, D.S., et al., CpG oligodeoxynucleotide- enhanced humoral immune response and production of antibodies to prion protein PrPSc in mice immunized with 139A scrapie- associated fibrils. J Leukoc Biol, 2007. 81(6): pp. 1374–1385.
Tayebi, M., Collinge, J., and Hawke, S., Unswitched immunoglobulin M response prolongs mouse survival in prion disease. J Gen Virol, 2009. 90(Pt 3): pp. 777–782.
Krasemann, S., et al., Induction of antibodies against human prion proteins( PrP) by DNA- mediated immunization of PrP0/ 0 mice. J Immunol Methods, 1996. 199(2): pp. 109–118.
Fernandez-Borges, N., et al., DNA vaccination can break immunological tolerance to PrP in wild- type mice and attenuates prion disease after intracerebral challenge. J Virol, 2006. 80(20): pp. 9970–9976.
Nitschke, C., et al., Immunisation strategies against prion diseases: prime- boost immunisation with a PrP DNA vaccine containing foreign helper T- cell epitopes does not prevent mouse scrapie. Vet Microbiol, 2007. 123(4): pp. 367–376.
Alexandrenne, C., et al., Electrotransfer of cDNA coding for a heterologous prion protein generates autoantibodies against native murine prion protein in wild- type mice. DNA Cell Biol, 2010. 29(3): pp. 121–131.
Nikles, D., et al., Circumventing tolerance to the prion protein( PrP): vaccination with PrP- displaying retrovirus particles induces humoral immune responses against the native form of cellular PrP. J Virol, 2005. 79(7): pp. 4033–4042.
Handisurya, A., et al., Vaccination with prion peptide- displaying papillomavirus- like particles induces autoantibodies to normal prion protein that interfere with pathologic prion protein production in infected cells. FEBS J, 2007. 274(7): pp. 1747–1758.
Rosset, M.B., et al., Dendritic cell- mediated- immunization with xenogenic PrP and adenoviral vectors breaks tolerance and prolongs mice survival against experimental scrapie. PLoS One, 2009. 4(3): p. e4917.
Goni, F., et al., Mucosal vaccination delays or prevents prion infection via an oral route. Neuroscience, 2005. 133(2): pp. 413–421.
Goni, F., et al., Mucosal immunization with an attenuated Salmonella vaccine partially protects white- tailed deer from chronic wasting disease. Vaccine, 2015. 33(5): pp. 726–733.
Goni, F., et al., High titers of mucosal and systemic anti- PrP antibodies abrogate oral prion infection in mucosal- vaccinated mice. Neuroscience, 2008. 153(3): pp. 679–686.
Bade, S., et al., Intranasal immunization of Balb/ c mice against prion protein attenuates orally acquired transmissible spongiform encephalopathy. Vaccine, 2006. 24(9): pp. 1242–1253.
Sacquin, A., et al., Prolongation of prion disease- associated symptomatic phase relates to CD3+ T cell recruitment into the CNS in murine scrapie- infected mice. Brain Behav Immun, 2012. 26(6): pp. 919–930.
Sadowski, M.J., et al., Anti- PrP Mab 6D11 suppresses PrP( Sc) replication in prion infected myeloid precursor line FDC- P1/ 22L and in the lymphoreticular system in vivo. Neurobiol Dis, 2009. 34(2): pp. 267–278.
Petsch, B., et al., Biological effects and use of PrPSc- and PrP- specific antibodies generated by immunization with purified full- length native mouse prions. J Virol, 2011. 85(9): pp. 4538–4546.
Didonna, A., et al., Characterization of four new monoclonal antibodies against the distal N- terminal region of PrP( c). PeerJ, 2015. 3: p. e811.
Pilon, J.L., et al., Immunization with a synthetic peptide vaccine fails to protect mule deer( Odocoileus hemionus) from chronic wasting disease. J Wildl Dis, 2013. 49(3): pp. 694–698.
David, M.A., Jones, D.R., and Tayebi, M., Potential candidate camelid antibodies for the treatment of protein- misfolding diseases. J Neuroimmunol, 2014. 272(1–2): pp. 76–85.
Han, Y., et al., Immune responses in wild- type mice against prion proteins induced using a DNA prime- protein boost strategy. Biomed Environ Sci, 2011. 24(5): pp. 523–529.