Recombinant Human Interleukin-11 (rHIL-11) Engineering Service

Recombinant Human Interleukin-11 (rHIL-11, Oprelvekin) is a crucial cytokine used to prevent severe thrombocytopenia in patients receiving chemotherapy. As a complex glycoprotein, it is typically produced in Chinese Hamster Ovary (CHO) cells. Production faces challenges due to its complex structure: low expression level in CHO cells results from metabolic limitations and transcription bottlenecks, leading to poor productivity. Furthermore, the protein is prone to degradation and high protein aggregation rate during purification and storage, drastically reducing the yield of the functional monomeric protein and hindering clinical formulation.

CD Biosynsis offers a comprehensive service focused on enhancing both the productivity and stability of rHIL-11 in a commercial context. Our core strategy involves optimization of fed-batch fermentation process in CHO cells . We employ sophisticated nutrient feeding strategies (e.g., concentrated glucose, amino acids, and specific precursors) combined with precise control of pH, dissolved oxygen, and temperature to prolong the high-viability production phase, boosting the overall volumetric yield. This is coupled with modification of protein surface charge . We utilize site-directed mutagenesis to introduce targeted substitutions of hydrophilic or charged residues on the protein surface. This modification aims to increase electrostatic repulsion and enhance hydration , thereby reducing the hydrophobic interactions responsible for protein aggregation without compromising the functional binding site. This integrated approach aims to deliver higher titers of a more stable, non-aggregated rHIL-11 product, improving both manufacturability and clinical efficacy.

Pain Points

Efficient and stable rHIL-11 production faces these key challenges in biomanufacturing:

• Low Expression Level in CHO Cells: Despite being the preferred host for glycosylation, CHO cells often struggle with low specific productivity (qP) due to nutrient limitations, waste accumulation, and sub-optimal transcription/translation rates.
• High Protein Aggregation Rate: rHIL-11 has a tendency to form inactive high-molecular-weight aggregates, especially under stress (high concentration, temperature shifts, shear stress). This leads to significant product loss during purification and formulation.
• Glycosylation Heterogeneity: The N-linked glycosylation of rHIL-11 can be variable in CHO cells, affecting its biological activity and pharmacokinetics .
• Yield/Viability Trade-off: Prolonging the production phase in fed-batch culture often results in decreasing cell viability and increasing protease activity , which further contributes to protein degradation.

A successful solution must simultaneously boost cellular productivity and ensure the long-term structural integrity of the secreted protein.

Solutions

CD Biosynsis utilizes advanced process and protein engineering to enhance rHIL-11 production in CHO cells:

Optimization of Fed-batch Fermentation Process in CHO Cells

           

We develop customized, chemically defined feed media (perfusion or bolus feeding) and optimize pH and temperature shifts to maximize integral viable cell concentration (IVCC) and specific productivity (qP).

Modification of Protein Surface Charge

We use site-directed mutagenesis to introduce strategically placed charged residues (e.g., Glu or Asp) on the surface to increase surface hydrophilicity and electrostatic repulsion , minimizing aggregation.

Host Cell Line and Vector Optimization

We screen different high-expressing CHO clones and use gene amplification systems (e.g., DHFR}/\text{MTX system) and optimized vector elements to enhance stable, high-level gene expression.

Aggregation Monitoring and Control

We implement online or rapid HPLC monitoring of aggregation during the run and use media supplements (e.g., stabilizing agents, specific surfactants) to reduce aggregation in the harvest fluid.

This systematic approach addresses both upstream (titer) and downstream (stability/aggregation) challenges simultaneously.

Advantages

Our rHIL-11 engineering service is dedicated to pursuing the following production goals:

Significantly Increased Titer

Process optimization leads to a higher viable cell density and prolonged production phase , boosting overall volumetric yield.

Reduced Aggregation and Enhanced Stability

Surface charge modification improves the structural integrity of rHIL-11, reducing product loss during manufacture and storage.

Cost-Efficient Manufacture

Higher yield and purity directly reduce the cost of goods (COG) , making the therapeutic more accessible. [Image of Cost Reduction Icon]

Consistent Glycosylation Profile

Process control (e.g., temperature) helps ensure a reproducible and optimal N-glycosylation profile for maximum activity.

High Bioactivity Retention

Protein engineering is designed to maintain or enhance receptor binding affinity while improving stability.

We provide a specialized platform focused on developing a high-performance, stable rHIL-11 production system.

Process

Our rHIL-11 engineering service follows a rigorous, multi-stage research workflow:

• Protein Mutagenesis and Cloning: Design and create rHIL-11 variants with surface charge modifications (e.g., targeted Lys-to-Glu substitutions) and clone into the CHO expression vector.
• Fed-batch Process Development: Conduct DoE (Design of Experiments) to optimize feeding strategies (timing and concentration) of glucose, glutamine, and trace elements to maximize IVCC.
• Cell Line Selection and Screening: Transfect and screen high-expressing CHO clones (e.g., CHO-K1 or CHO-S), followed by limiting dilution cloning for monoclonality.
• Aggregation Analysis and Formulation: Use analytical tools (e.g., SEC-HPLC, DLS) to characterize aggregation levels in the harvest fluid and test different media supplements/buffers for stability.
• Scale-up and Bioreactor Validation: Perform demonstration runs in bench-scale bioreactors to confirm the titer, viability, and aggregation reduction of the optimized clone and process.
• Result Report Output: Compile a detailed Experimental Report including cell line data, fed-batch parameters, and functional data (titer, aggregation %, bioactivity) , supporting cGMP transfer.

Technical communication is maintained throughout the process, focusing on timely feedback regarding expression titer and stability.

Explore the potential for a high-titer, high-stability rHIL-11 therapeutic. CD Biosynsis provides customized cell line and process engineering solutions:

• Detailed Stability and Aggregation Report , demonstrating the reduction of high-molecular-weight species post-mutation and process optimization.
• Consultation on optimized CHO cell culture media and feed formulation for improved IVCC.
• Experimental reports include complete raw data on volumetric titer (g/L) and specific productivity (qP) , essential for process economics.