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Institute of Molecular Biology at the Bulgarian Academy of Sciences, Sofia, Bulgaria

This labs research is focused on the quality of recombinant proteins produced in Escherichia coli. They are searching for the reasons causing structural instability and immunogenicity of therapeutic proteins and for ways how to overcome these problems. They discovered an unknown cause of protein drugs’ immunogenicity – the involvement of the proteins in the Maillard reaction (glycation) during their synthesis in the bacterial host. The Maillard reaction results in the formation of sugar-derived neo-epitopes in the proteins known as advanced glycation end products (AGEs). The lab further explores means to avoid or at least minimize glycation of the recombinant proteins throughout the manufacturing process starting with fermentation and ending up with drugs’ formulation and storage.

Diseases Researched:

They are developing protein drugs for treatment of:

  • Viral diseases
  • Autoimmune diseases

They have developed two pharmaceutical formulations based on human γ-interferon (hIFNγ):

  • Eye drops for treatment of eyes’ viral infections (Gammaferon)
  • Gel (Virogel) for treatment of Herpes zoster

Current projects are committed to the development of protein drugs for management of autoimmune disorders linked to the endogenous overproduction of hIFNγ such as:

  • Multiple sclerosis
  • Alopecia areata
  • Myasthenia gravis
  • Autoimmune uveitis

Performed Tests:

Drugs measured:

hIFN-γ drugs

  • Kynurenine assay (with modification)- Measurement of the biological activity of hIFNγ
  • Sandwich and direct ELISA- Measurement of hIFNγ concentration
  • Isothermal titration calorimetry (ITC)- Measurement of drug-target (hIFNγ-hIFNγR) interactions

ADAs measured:

hIFN-β binding and neutralizing antibodies, cross-reactivity of anti-IFNβ antibodies with advanced glycation end products (AGEs)

  • Measurement of binding anti-IFN-β and anti-AGEs antibodies with direct ELISAs
  • Measurement of the cross-reactivity of anti-IFN-β antibodies with AGEs with competitive ELISA
  • Using commercial hIFNβ1a and their own screening protocol to measure binding antibodies in human sera with indirect ELISA
  • Measurement of hIFNβ1a neutralizing antibodies in human sera with Functional immunoassay with commercial iLite Type I IFN Assay Ready Cells

Website link: http://www.bio21.bas.bg/imb/?id=3