Available Technology
Bacteria based PrP and Amyloid technology generates novel aggregation assays
Technology:
Assay for identifying amyloidogenic proteins
Markets Addressed
Abnormal aggregation and accumulation of amyloid-forming proteins can lead to a variety of protein aggregation disorders known as amyloidoses, including type 2 diabetes, Alzheimer’s (AD), Parkinson’s (PD), Huntington’s (HD) and prion diseases. To date the mechanistic basis of prion-mediated cytotoxicity remains poorly understood. Despite 40 years of intensive scientific attention, no effective treatments have been developed for controlling mammalian prion diseases.
Underlying these inevitably fatal neurodegenerative diseases is the specific cellular protein PrP, which has the potential to form self-propagating aggregates that are infectious. These aggregates are composed of highly structured sheet-rich fibrils known as amyloids, and conversion to the fibrillar form (PrPres) involves a specific change in the conformation of PrP.
Two important factors continue to hinder the development of therapeutics for Prion disease:
1. The mechanistic basis of prion-mediated cytotoxicity remains poorly understood. Although the majority of studies aimed at uncovering therapeutics for prion disease have targeted PrPres, an increasing body of evidence points to pre-fibrillar aggregates of PrP as the most toxic species, suggesting that it may be critical to target the earliest steps in the conversion of soluble PrP to PrPres.
2. Whereas many amyloidogenic proteins readily undergo conversion to the Amyloid form in vitro, the de novo conversion of purified recombinant PrP to the infectious, aggregated form has been accomplished only with great difficulty. Soluble PrP must typically be treated with denaturants to promote protein misfolding and subjected to multiple cycles of sonication and incubation (a procedure called Protein Misfolding Cyclic Amplification, or PMCA) in the presence of facilitating factors to amplify the aggregated form. The requirement for these cumbersome manipulations complicates efforts to study the misfolding events or the intermediate oligomeric species generated during PrP aggregation.
To circumvent some of these difficulties, alternative Amyloid-forming proteins are being explored as surrogate systems for the discovery of therapeutics for controlling prion diseases. The Hochschild lab proposed to implement an E. coli-based assay to identify small molecules that could modulate the intracellular aggregation of a bacterial Amyloid-forming protein. The lab had since developed a new E. coli-based assay that facilitates the efficient fibrillization of a variety of amyloidogenic proteins at the cell surface. This assay provides a new experimental approach for studying PrP.
Innovations and Advantages
Scientists at the Hochschild lab have directly exploited the ability of E. coli cells to assemble Amyloid fibers at the cell surface to develop a general assay for identifying amyloidogenic proteins.
The de novo conversion of purified recombinant PrP to the infectious, aggregated form has been accomplished only recently and using a lengthy and cumbersome procedure. The proposed E. coli-based system has the potential to provide a greatly simplified means to study the PrP conversion process and the effects of facilitating factors and other prospective modulators of PrP conversion and aggregation.
This technology could be utilized for two different assays:
1. A cell growth assay to screen small molecules that can modulate the intracellular aggregation of a bacterial Amyloid-forming protein.
This technology can be adapted for high-throughput screens to detect small molecules that can disrupt the assembly and/or promote the clearance of the toxic aggregates.
2. An assay for monitoring the abilities of specific target proteins to form Amyloid aggregates.
This could serve as a platform for the mass production of research tools to study conversion processes of specific Amyloid-forming proteins and the effects of modulators of Amyloid conversion and aggregation. This assay may also be adapted to a sensitive method to detect infectious materials.
This technology makes it possible to study or identify compounds that target the intermediate process towards Amyloid aggregation including the initial formation of soluble oligomers. Finally, it is fast, inexpensive, and highly reproducible. The technology is highly versatile and can be used to study a broad range of Amyloid-forming protein and their derivatives.
Additional Information
Intellectual Property Status: Patent(s) pending
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Inventor(s):
Hochschild, Ann
Sivanathan, Viknesh
Categories:
For further information, please contact:
Michal Preminger, Director of Business Development
(617) 432-0920
Reference Harvard Case #4406