Erratum: A tetracationic porphyrin with dual anti-prion activity.

[This corrects the article DOI: 10.1016/j.isci.2023.107480.].

Understanding the key features of the spontaneous formation of bona fide prions through a novel methodology that enables their swift and consistent generation.

Among transmissible spongiform encephalopathies or prion diseases affecting humans, sporadic forms such as sporadic Creutzfeldt-Jakob disease are the vast majority. Unlike genetic or acquired forms of the disease, these idiopathic forms occur seemingly due to a random event of spontaneous misfolding of the cellular PrP (PrPC) into the pathogenic isoform (PrPSc). Currently, the molecular mechanisms that trigger and drive this event, which occurs in approximately one individual per million each year, remain completely unknown. Modelling this phenomenon in experimental settings is highly challenging due to its sporadic and rare occurrence. Previous attempts to model spontaneous prion misfolding in vitro have not been fully successful, as the spontaneous formation of prions is infrequent and stochastic, hindering the systematic study of the phenomenon. In this study, we present the first method that consistently induces spontaneous misfolding of recombinant PrP into bona fide prions within hours, providing unprecedented possibilities to investigate the mechanisms underlying sporadic prionopathies. By fine-tuning the Protein Misfolding Shaking Amplification method, which was initially developed to propagate recombinant prions, we have created a methodology that consistently produces spontaneously misfolded recombinant prions in 100% of the cases. Furthermore, this method gives rise to distinct strains and reveals the critical influence of charged surfaces in this process.

A tetracationic porphyrin with dual anti-prion activity.

Prions are deadly infectious agents made of PrPSc, a misfolded variant of the cellular prion protein (PrPC) which self-propagates by inducing misfolding of native PrPC. PrPSc can adopt different pathogenic conformations (prion strains), which can be resistant to potential drugs, or acquire drug resistance, hampering the development of effective therapies. We identified Zn(II)-BnPyP, a tetracationic porphyrin that binds to distinct domains of native PrPC, eliciting a dual anti-prion effect. Zn(II)-BnPyP binding to a C-terminal pocket destabilizes the native PrPC fold, hindering conversion to PrPSc; Zn(II)-BnPyP binding to the flexible N-terminal tail disrupts N- to C-terminal interactions, triggering PrPC endocytosis and lysosomal degradation, thus reducing the substrate for PrPSc generation. Zn(II)-BnPyP inhibits propagation of different prion strains in vitro, in neuronal cells and organotypic brain cultures. These results identify a PrPC-targeting compound with an unprecedented dual mechanism of action which might be exploited to achieve anti-prion effects without engendering drug resistance.

Bona fide atypical scrapie faithfully reproduced for the first time in a rodent model.

Atypical Scrapie, which is not linked to epidemics, is assumed to be an idiopathic spontaneous prion disease in small ruminants. Therefore, its occurrence is unlikely to be controlled through selective breeding or other strategies as it is done for classical scrapie outbreaks. Its spontaneous nature and its sporadic incidence worldwide is reminiscent of the incidence of idiopathic spontaneous prion diseases in humans, which account for more than 85% of the cases in humans. Hence, developing animal models that consistently reproduce this phenomenon of spontaneous PrP misfolding, is of importance to study the pathobiology of idiopathic spontaneous prion disorders. Transgenic mice overexpressing sheep PrPC with I112 polymorphism (TgShI112, 1-2 × PrP levels compared to sheep brain) manifest clinical signs of a spongiform encephalopathy spontaneously as early as 380 days of age. The brains of these animals show the neuropathological hallmarks of prion disease and biochemical analyses of the misfolded prion protein show a ladder-like PrPres pattern with a predominant 7-10 kDa band. Brain homogenates from spontaneously diseased transgenic mice were inoculated in several models to assess their transmissibility and characterize the prion strain generated: TgShI112 (ovine I112 ARQ PrPC), Tg338 (ovine VRQ PrPC), Tg501 (ovine ARQ PrPC), Tg340 (human M129 PrPC), Tg361 (human V129 PrPC), TgVole (bank vole I109 PrPC), bank vole (I109I PrPC), and sheep (AHQ/ARR and AHQ/AHQ churra-tensina breeds). Our analysis of the results of these bioassays concludes that the strain generated in this model is indistinguishable to that causing atypical scrapie (Nor98). Thus, we present the first faithful model for a bona fide, transmissible, ovine, atypical scrapie prion disease.

Pharmacological inactivation of the prion protein by targeting a folding intermediate.

Recent computational advancements in the simulation of biochemical processes allow investigating the mechanisms involved in protein regulation with realistic physics-based models, at an atomistic level of resolution. These techniques allowed us to design a drug discovery approach, named Pharmacological Protein Inactivation by Folding Intermediate Targeting (PPI-FIT), based on the rationale of negatively regulating protein levels by targeting folding intermediates. Here, PPI-FIT was tested for the first time on the cellular prion protein (PrP), a cell surface glycoprotein playing a key role in fatal and transmissible neurodegenerative pathologies known as prion diseases. We predicted the all-atom structure of an intermediate appearing along the folding pathway of PrP and identified four different small molecule ligands for this conformer, all capable of selectively lowering the load of the protein by promoting its degradation. Our data support the notion that the level of target proteins could be modulated by acting on their folding pathways, implying a previously unappreciated role for folding intermediates in the biological regulation of protein expression.