Amal Abukhares

Pharmaceutical Conferences-Amal Abukhares

Title: A Pre-formulation study of recombinant human interleukins-1 receptor antagonist for emergency use

Amal Abukhares

University of Manchester, UK


Amal obtained her first degree in Pharmacy from the University Of Tripoli, Libya 2004-2005. In 2006 she starts to work at University of Tripoli as teaching assistant (demonstrator).  She is involved in the supervision of undergraduate students in the assistance for the smooth running of the laboratories. In 2007 she has received funding from her University to finish her post-graduation study, under Libyan grant agreement of 701/2007. In September 2010 she completed her Master degree in drug delivery from Aston University/UK. Between 2011 and 2014 she worked as lecturer assistant in industrial pharmacy department at Pharmacy School of University of Tripoli. She gave lectures and run lab works. She has received another funding from her the University on 2013 under Libyan grant agreement of 293/2013.In Jan 2015 she joined the University of Manchester for a PhD in Pharmaceutical sciences. Her role as PhD student is to conduct research to evaluate and improve stability of a protein called rhIL-1Ra. Find out root causes of instability of rhIL-1Ra. Conduct several pre-formulation studies by  using different biophysical techniques such as Static and Dynamic Light Scattering (OD,DLS), Circular Dichroism (CD), Differential Scanning Colorimeter DSC and SDS-Electrophoresis. Find a new formulation technique to address stability problem of the protein.  Currently, she is also work as teaching assistant (demonstrator) at University of Manchester.


Introduction: rhIL-1Ra is a recombinant form of the natural anti-inflammatory mediator, interleukin-1 receptor antagonist that competitively binds with type I receptors (IL-1RI). It inhibits effects of other inflammatory mediator, interleukin -1 (IL-1). The product is licensed on 2013 to treat Rheumatoid Arthritis. There is preclinical evidence to suggest that IL-1Ra can be a promising candidate to treat brain stroke. Treatment with Kineret® results in reduced levels of inflammatory markers in plasma of stroke patients and the cerebrospinal fluid and plasma of patients with subarachnoid haemorrhage. However the Kineret® has stability problems at ambient temperature and is hence stored at 2°C to 8°C. In addition, the manufacturer (Amgen PLC) recommends avoiding shaking, freezing and exposure to sunlight. These recommendations for storage are incompatible if the formulation is to be used treat brain stroke outside hospital settings, particularly by paramedic staff in ambulances. It would be advantageous to treat stroke rapidly through intervention by paramedics. This would minimise brain damage from the first sign of stroke.

Aim: The overall project aim is to study thermal stability of the IL-1Ra and identify the root cause of instability. This would lead to designing a suitable pre-formulation to address concerns with storage at ambient temperature.

Methods: Initially, visual inspection (photographs) was made following the incubation of Kineret® at 37°C. Then, the aggregation was followed quantitatively by recording changes in the optical density at 450 nm (OD450 nm). A more sensitive technique was also used to follow aggregation in the nucleation phase, where sub-visible particles may exist, using Dynamic light scattering (DLS). The formation of non-native β-structure of the protein was followed by use fluorescence spectroscopy using Thioflavin T (THT). Attempts were also made, using circular dichroism (CD), to assess any induced changes in the secondary structure of the protein induced thermally.

Conclusion: Our study from suggests that formulation shows visible signs of cloudiness within 6hrs at 37° C. When this is followed by light scattering over 24hrs period a number of kinetic phases due to aggregation were observed starting with a nucleation phase where the changes in particle size were confirmed by DLS. Protein content show a trace amount of protein was aggregated. Fluorescence assay in particular the Thioflavin T result provides earlier indication of formation of non-native beta structures. Therefore, further study will be conducted to characterise protein aggregates