Prions: Protein Aggregation and Infectious Diseases - Physiological ...

1138 ADRIANO AGUZZI AND ANNA MARIA CALELLA
tive value of the different in vitro and in vivo experiments
is limited, and further investigations will be required.
C. Soluble Prion Antagonists
In several paradigms, expression of two PrP C moieties
with subtle differences antagonizes prion replication.
The molecular basis for these effects is unknown.
Perhaps the subtly modified PrP C acts as a decoy by
binding incoming PrP Sc (or protein X) and sequestering it
into a complex incapable of further replication.
To test the latter hypothesis, a transgenic mouse
was developed that expresses soluble full-length mouse
PrP rendered dimeric by fusion with the Fc portion of
human IgG1 (known as PrP-Fc 2) (Fig. 10D). After prion
inoculation, these mice were surprisingly resistant to
prion disease (343). The PrP-Fc 2 was not converted to
a prion-disease-causing isoform. Moreover, when the
transgenic mice expressing PrP-Fc 2 were back-crossed
with wild-type mice and then inoculated with prions,
they showed marked retardation in the development of
prion disease, which was equivalent to a 10 5 -fold reduction
in titer of the prion-infected inoculum. This antiprion
effect occurred in two different lines of PrP-Fc 2expressing
transgenic mice after either intracerebral or
intraperitoneal infection with scrapie. Therefore, it
seems that PrP-Fc 2 effectively antagonizes prion accumulation
in the spleen and brain. Because PrP-Fc 2 cannot
be converted to the protease-resistant, diseasecausing
isoform, it might be effectively functioning as a
“sink” for PrP Sc , by binding PrP Sc and preventing the
binding and conversion of PrP C (20).
Delivery of the soluble prion antagonist PrP-Fc 2 to
the brains of mice by lentiviral gene transfer impaired
replication of disease-associated PrP Sc and delayed disease
progression (178). These results suggest that somatic
gene transfer of prion antagonists may be effective for
postexposure prophylaxis of prion diseases. In addition, it
remains to be established whether the current form of
PrP-Fc 2 has the strongest antiprion properties. For instance,
the introduction of dominant-negative mutations
analogous to those described for Prnp (374, 391) might
considerably augment its efficacy. However, further research
is needed to establish whether it may be effective
as a biopharmaceutical.
IX. PROGRESS IN THE DIAGNOSTICS OF
PRION DISEASES
As with any other disease, early diagnosis would
significantly advance the chances of success of any possible
intervention approaches. Unfortunately, prion diagnostics
continue to be rather primitive. Presymptomatic
diagnosis is virtually impossible, and the earliest possible
Physiol Rev VOL 89 OCTOBER 2009 www.prv.org
diagnosis is based on clinical signs and symptoms. Hence,
prion infection is typically diagnosed after the disease has
already progressed considerably.
A significant advance in prion diagnostics was accomplished
in 1997 by the discovery that proteaseresistant
PrP Sc can be detected in tonsillar tissue of
vCJD patients (221). It was hence proposed that tonsil
biopsy may be the method of choice for diagnosis of
vCJD (218). Furthermore, there have been reports of
individual cases showing detectable amounts of PrP Sc
at preclinical stages of the disease in the tonsil (443) as
well as in the appendix (223), indicating that lymphoid
tissue biopsy may represent a potential test for asymptomatic
individuals. These observations triggered large
screenings of human populations for subclinical vCJD
prevalence, using appendectomy and tonsillectomy
specimens (187). PrP Sc -positive lymphoid tissues were
long considered to be a vCJD-specific feature that
would not apply to any other forms of human prion
diseases (218). However, a successive survey of peripheral
tissues of patients with sporadic CJD has led to the
identification of PrP Sc in as many as one-third of skeletal
muscle and spleen samples (183). In addition, PrP Sc
was found in the olfactory epithelium of patients suffering
from sCJD (532). These unexpected findings
raise the hope that minimally invasive diagnostic procedures
may take the place of brain biopsy in intravital
CJD diagnostics.
For almost three decades, all gold standard methods
for the molecular diagnosis of prion diseases have relied
on the use of proteinase K (PK) to differentiate PrP C and
PrP Sc . Recently, the complementary use of the protease
thermolysin has been introduced. This protease digests
PrP C but, unlike PK, leaves PrP Sc intact without truncation
of the NH 2 terminus (126, 376).
The development of highly sensitive assays for biochemical
detection of PrP Sc in tissues and body fluids is a
top priority. One way to achieve this goal is to develop
high-affinity immunoreagents that recognize PrP Sc . Examples
include the “POM” series of antibodies that recognize
various well-defined conformational epitopes in the structured
COOH-terminal region of PrP C , and linear epitopes
in the unstructured NH 2-terminal region (395). Because of
the particular nature of the epitopes to which they are
directed, some of these antibodies have affinities for the
prion protein in the femtomolar range. Antibodies that
specifically bind PrP Sc without binding PrP C have also
been reported (279, 358), yet their affinity seems to be
limited and their diagnostic value has awaited confirmation
for more than one decade to no avail.
Searching for PrP Sc binding reagents, Lau et al. (292)
discovered PrP-derived peptides that bound PrP Sc . When
coupled with a sandwich ELISA for detection, these peptide
binding reagents create a sensitive assay that can detect
PrP Sc nanoliter amounts of 10% (wt/vol) vCJD brain homog-