Prothyx Technology Platform Brief
A novel drug delivery approach conferring latency, stability and disease targeting benefits to protein-based therapeutics

Background
Stealthyx Therapeutics Ltd was formed to exploit the novel Prothyx technology platform that was developed in the laboratory of Prof. Yuti Chernajovsky at Barts and The London, Queen Mary's School of Medicine and Dentistry, University of London.

Prothyx is a molecular biology platform that harnesses the properties of the naturally occurring peptide LAP to confer stability, latency and disease targeting to protein-based therapeutic macromolecules. Activation and release of the therapeutic entities occurs in a disease-specific manner via proteolytic cleavage. The approach allows for the production of biological therapeutics that exhibit improved performance characteristics and provide competitive benefits to protein-based therapeutics.

Application areas
Prothyx can be applied to a variety of protein and related therapeutic compounds, including cytokines, growth factors, pro-drug converting enzymes and scavenging enzymes, including molecules that would otherwise show systemic toxicity. Future developments will seek to expand the technology to other classes of drug candidates, including protein nucleic acids (PNA) and synthetic compounds.

Prothyx-modified drugs are compatible with diseases that involve inflammation, such as many autoimmune conditions, and tissue remodelling, such as cancer, osteoporosis, atherosclerosis etc. In addition, the platform may be applicable to certain infectious diseases.

Application Benefits
The Prothyx system has the potential to confer the following clinical benefits to a biological therapy:

Increased therapeutic index: higher doses and fewer observable side effects because of the latency conferred by the Prothyx platform. Activation and release of the therapeutic component occurs primarily at the site of disease.

Extended stability in the circulation: latent Prothyx-modified protein therapeutics have a prolonged half-life in the circulation, allowing longer intervals between parenteral administrations and improving patient compliance.

Prothyx functionality: latency, stability and disease targeting
Prothyx technology enables fusion molecules to be created between human LAP and proteins of therapeutic potential. This fusion confers both enhanced stability and latency to the therapeutic moiety by providing a 'shell' structure surrounding the active therapeutic component.

Schematic illustration of the Prothyx platform

Active therapeutic molecules are released from the Prothyx shell by the action of enzymes present at the sites of disease, such as the family of matrix metalloproteases. Disease-specific activation and release is engineered into the drug candidates by the creation of selective cleavable linkers between the LAP and therapeutic components. Modified from Adams et al. Nature Biotechnology 21:1314-1320; 2003.

Scientific Rationale: the LAP peptide
LAP is a naturally occurring peptide that is expressed as a 'leader' sequence of newly synthesized molecules of the cytokine TGF-Beta. The peptide remains associated with TGF-Beta, forming an inactive and stable reservoir from which active cytokine is released through the action of a number of physiological stimuli. The Prothyx platform exploits these natural properties of LAP to derive protein therapeutics with enhanced performance characteristics.

Exemplification of Prothyx
Initial exemplification work has been undertaken using Interferon-Beta, other cytokines and peptides as the therapeutic components.

Stealthyx has demonstrated:
Expression of functional LAP fusion molecules: Prothyx-modified molecules incorporating a number of different cytokines and peptides have been successfully expressed in mammalian cells.

Purification of LAP fusion proteins: A generic single-step method for the purification of Prothyx molecules has been developed.

Development of disease-specific cleavable linkers: Cleavable linkers with selectivity for fluids from disease sites and showing different protease sensitivities have been developed.

Demonstration of prolonged stability of LAP fusion molecules: Parenteral administration of Prothyx-modified IFN showed significantly extended half life in the circulation of rodents.

Evidence of in vivo efficacy: Prothyx -modified IFN delivered by DNA intramuscular injection showed increased therapeutic efficacy after disease onset compared to unmodified IFN in a collagen-induced arthritis mouse model.

Accessing the Prothyx Platform
Stealthyx has developed plasmids, cleavable linker sequences, mammalian expression vectors, purification protocols and assays to enable production and evaluation of Prothyx-modified candidate molecules.

Current Status
Stealthyx Therapeutics Ltd is seeking discovery partnerships with R&D organisations to co-develop Prothyx with their biological therapies and license the technology for drug development.
Stealthyx is continuing to characterise the Prothyx system using IFN-b as a treatment for autoimmune diseases and as a delivery vehicle for other cytokines and cytotoxic compounds. The Company is also undertaking detailed animal studies to characterise the in vivo benefits of the Prothyx system.

For Further Information
To discuss potential collaborations with Stealthyx Therapeutics Ltd, please contact:

Research:
Professor Yuti Chernajovsky
Stealthyx Therapeutics Ltd
Bone and Joint Research Unit
Barts and The London
Queen Mary's School of Medicine and Dentistry, University of London
Charterhouse Square
London, EC1M 6BQ

Email: yuti.chernajovsky@stealthyx.com
Tel: +44 (0)207 882 6122

Business Development, acting on behalf of Stealthyx Therapeutics Ltd:

c/o Donna Hackett
Partner
Commercial and Legal
ProPharma Partners Limited
10, The Courtyard
East Park
Crawley
West Sussex
RH10 6AG

M: +44 (0) 771 423 4705
F: +44 (0) 1293 415893
E: dhackett@propharmapartners.uk.com