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  • 80+ CMC & CMO Projects

  • 20 Global IND Approvals

By Dec.2022

Progress of Typical Projects

Client Type Stage
Leading biopharma in China Recombinant Protein Vaccine NMPA IND clearance
Clinical phase 3
KLT Cytokine NMPA IND clearance
Clinical phase 2
Biotech in China Trispecific antibody NMPA IND clearance
FDA IND Clearance
Clinical phase 1
Biotech in Korea Ab-cytokine conjugated FDA IND clearance
Clinical phase 1
Biotech in China Bispecific antibody NMPA IND filing

By July, 2022

Cell Line Development
Upstream Process
Downstream Process
Analytical Development
  • Focus on product quality
  • Titer improvement
  • Optimize product quality based on molecule features
  • High throughput parameter screening
  • Ion additive
  • Harvesting time
  • Filter adsorption
  • Resin selection
  • S/D inactivation
  • PTM
  • Analytical method development capability
  • Special assay kit development

Cell Line Development—Focus on quality and titer

GenScript ProBio’s cell line development platform is based on the proprietary CHOK1-GenS system with high productivity and excellent stability. The average titer can reach 4.35g/L. CHOK1-GenS system is compatible with different molecular types, such as bispecific antibody and recombinant protein. It only takes less than 3 months from DNA to PCB.

For recombinant protein, GenScript ProBio pays attention to product quality and purity, and increase cell pool and clone selection scale to improve final titer.

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Upstream Process Development

1. Through the use of high-throughput platforms such as Ambr15 for process development, GenScript ProBio can efficiently complete clone screening, media screening and parameter optimization (i.e. pH, temperature, feed strategy). ProBio optimizes product quality based on molecular features.

  • 2. Suitable additives to improve product quality
    In a recombinant protein case, ion additives help improve monomer purity.

  • 3. Define the best harvesting time
    By extending the cell culture time and analyzing the trend of activity and viability, ProBio can define the best harvesting time is D11.

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Downstream Process Development

  • 1.Attention to filter adsorption
    Variant filter adsorption in recombinant protein cases.

    C filter shows the lowest adsorption performance.

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  • 2. S/D substitutes low pH
    Protein will fully precipitate under low pH, while S/D inactivation method can act as the substitution. S/D can also help improve the purity due to the aggregate precipitation.

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  • 3. AC resin selection
    Many recombinant proteins are not suitable for protein A affinity chromatography. Multimodal resin can fulfill the purity and yield requirements.

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  • 4. Polish resin selection
    Multimodal resin is suitable for recombinant protein polish purification and can remove aggregates.

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Analytical Development

Recombinant protein has special molecular properties. Conventional analysis methods cannot meet the needs of the projects. So recombinant protein projects need strong analytical development platform, including structure, residuals and bioactivity. This table shows some challenges and solutions in the protein projects.

Challenge Solution
Recombinant protein is unstable in mobile phase
Standard mAb SEC is not applicable
Mobile phase optimization and selection
Recombinant protein may precipitate during pretreatment
Standard mAb glycan analysis is not applicable
Optimize pretreatment buffer
Increase sample volume
O-glycan site identification
ESI-MS is not applicable
SialEXO&OpeRATOR
LC-MSMS to identify the O-glycan site
O-glycan quantitative analysis
β-elimination method is not stable
LC-MSMS measurement
No commercial Chaperon assay kit
No established method
Develop Chaperon assay kit based on ELISA

Case Study: Recombinant Protein

Entered into clinical phase Ⅱ

Content Customer Expectation GenScript Delivered Note
Titer 30 mg/L 500 mg/L Titer improved 15 times
Pilot scale only need 7L
Pilot Scale 100L 7L
Purity SDS-PAGE ≥95% >98% Purity >98% by USP&DSP
SEC-HPLC ≥95% >98%
RP-HPLC ≥95% >98%
Impurity Aggregate ≤2.0% <1.0% Aggregate <1.0%
P35 residue ≤1.0% Undetectable Remove p35, p40 and p80 residue effectively by DSP
P40 residue ≤1.0% Undetectable
P80 residue/td> ≤1.0% Undetectable
HCP ≤0.05% ≤0.01% Low level of host residual
HCD ≤10ng/Dose ≤0.01ng/Dose
In vitro activity 80%-150% 90%-110% Stable bioassay method
Consistent batch quality
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Cell Line Development—Focus on quality and titer

GenScript ProBio’s cell line development platform is based on the proprietary CHOK1-GenS system with high productivity and excellent stability. The average titer for mAb can reach 4.35g/L and the average titer for bsAb is 2.3g/L. It only takes less than 3 months from DNA to PCB.

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Upstream Process Development

  • 1. Suitable additives to improve product quality
    In one bispecific antibody case, modified amino acids help improve titer to 150%

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  • 2. Process optimization to improve titer
    In one bispecific antibody case, optimizing culture conditions such as pH and temperature to improve titer.

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  • 3. Process intensification
    In addition to the traditional fed-batch process, GenScript ProBio also provides process intensification solutions such as high density inoculation to help improve titer.

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  • 4. Perfusion culture helps improve product quality
    Perfusion culture helps increase full length ratio and purity

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Downstream Process Development

  • 1. High-throughput downstream process screening platform

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  • 2. S/D method in place of low pH can help improve purity

    VI Method Sample ID Conc.(mg/ml) SEC-HPLC(%) CE-SDS-N
    HMW Main LMW (%)
    Control AC Eluate 12.44 1.65 98.35 ND 99.61
    S/D AC Eluate 12.60 1.47 98.54 ND 99.54
    Low pH pH3.8, 1hr 10.73 3.33 96.42 0.24 99.11
    pH3.7, 1hr 10.39 14.61 85.40 ND 99.18
  • 3. Aggregate removal
    Multimodal resin is suitable for multispecific antibody purification and can remove aggregates.
    In one trispecific antibody case, through downstream process optimization, purity was improved from 84.7% to 98.6%.

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Analytical Development

Multispecific antibodies often have chain mismatches, so appropriate analysis methods need to be developed to monitor the occurrence of mismatches.

LC-MS method can sensitively monitor chain mismatch.

GenScript ProBio has strong analytical development capabilities, and can design suitable bioassay and functional activity methods based on molecule features and MOA

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