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Protein Formulation
Workflow
In-house Development Capability
Rational for Micro-scale, Parallel
Protein Formulation Workflow
“There is the need for more complex and orthogonal analytical
technologies to detect all of the chemical and physical instabilities that
may result for large molecules during formulation development”
> Parallel formulation development to increase experimental breadth
> Micro-scale systems to reduced drug substance requirements
> Automation to reduce human error and increase throughput
> Electronic data management to improve communication and analysis
> Reduce the barrier to screening to move formulation development earlier
into the product development process
Protein Formulation Workflow
Sample Conditioning Chambers
UV-Vis / Florescence Reader
& LabChip GEL SDS
Chromatography
Analytical Systems
Plate Stackers and
Label Applicators
Dynamic Light
Robotic Sample Scattering Station
Preparation and Analytical
Work-up stations
High Throughput
Protein Formulation Process
Automated Formulation System
Protein Solution (Sample volumes as
low as 125 µL) Agitation Testing
Preparation
For PhotoStability Studies
surfactant
Freeze Thaw Cycling
screening
Formulation/ DSC
Sample Preparation
Robot
Details: Final Formulation Environmental
Formulation Prep / Dialysis
Adjustment, Plate/Vial Chambers
Station
handle, Image Samples, Details: 4
Details: Prepare buffer exchange Analytical Sample Prep Temperatures, 4
solutions and perform dialysis with weights, pH time points
Analytical
Sterile Filtration / Filling / Stations
Capping Station UV-Vis, FI / SEC /
RPLC / IEC
Lyophilization GelElectrophoresis
Station Osmometer
High Throughput
Protein Formulation Process
Design Library
Prepare
Formulations
Stress Testing
Analytical
Sample Preparation
Sample Analysis
Data Processing,
Visualization
Report Generation
Protein Formulation Solubility Results
High-Throughput vs Traditional Manual-based Measurements
No Surfactant 0.05wt% P20
Protein Solubility versus pH with 150mM NaCl as Tonicity
Modifier
25
No Tonicity Modifier
20
•Manual Study
50mM NaCl (Multi-week)
Solubility (mg/mL)
15
•HT Approach Study
Manual Multi-Week
150mM NaCl High-Throughput Study
Recombinant Protein Concentration 10 (2-3 days)
17.8 .. 20.1 250mM NaCl
5
15.6 .. 17.8
13.4 .. 15.6
11.3 .. 13.4 2.5wt% Sorbitol
0
9.1 .. 11.3 pH 4 pH 5 pH 6 pH 7 pH 8
6.9 .. 9.1
4.7 .. 6.9 5.0wt% Sorbitol
2.5 .. 4.7
0.3 .. 2.5
1.5wt% Glycine Protein Solubility versus pH with 5% Sorbitol as Tonicity
Modifier
Units: mg/mL
3.0wt% Glycine 25
20 •Manual Study
(Multi-week
10mM Sodium Phosphate Buffer, pH 7
10mM Sodium Phosphate Buffer, pH 8
Solubility (mg/mL)
10mM Sodium Acetate Buffer, pH 4
10mM Sodium Acetate Buffer, pH 5
10mM Histidine Buffer, pH 6
10mM Sodium Phosphate Buffer, pH 7
10mM Sodium Phosphate Buffer, pH 8
10mM Histidine Buffer, pH 7
10mM Sodium Acetate Buffer, pH 4
10mM Sodium Acetate Buffer, pH 5
10mM Histidine Buffer, pH 6
10mM Histidine Buffer, pH 7
15
•HT Approach Study
Manual Multi-Week
10
(2-3 days)
High-Throughput Study
5
0
pH 4 pH 5 pH 6 pH 7 pH 8
Protein Formulation Surfactant Screen
DLS vs SEC-HPLC Results
Average Radius from Dynamic Light Scattering Size Exclusion Chromatography: Percent Area of Aggregates Peak
Protein A
P20
P80
F68
Summary
Workflow Functionality
• 4x more formulations with same amount of protein
– Formulation volumes as low as 125 µl
• Fully automated Analytical systems and Sample preparation
– HPLC ( Size Exclusion, Ion Exchange, Reversed Phase, HIC)
– UV-vis and Fluorescence Spectrometry.
– Dynamic Light Scattering
– LabChip Gel Electrophoresis
– Osmometer
• Fully Automated Sample/Equipment Tracking and Record Keeping
– Bar codes to track Vial racks
– Future: RFID tags/label to track mobile phases, columns, analytical equipment.
– Future: Ultrasound and Optical sensors for rack/vial/cap detection and error handling
Applications
• Solubility screens mapping short-term thermodynamic solubility for high concentration formulations
• Automated sample preparation and analysis for HPLC, DLS, Chip Electrophoresis, pH, osmometry on
microscale samples
• Agitation and freeze-thaw short-term tests (non aseptic) for stability assessment at the microscale
• Preformulation studies in final container-closure systems in an aseptic environment
• 12 to 48 different formulations (250mg to 1g of protein for the entire study)
• Stresses: temperature, photolytic, oxidative, agitation, freeze-thaw
• At least 4 time points for each sample at each stress condition
