Fusion of Enhanced Cell Performance with Improved Productivity:

Development of a Robust Chemically Defined Formulation for

Culture of Chinese Hamster Ovary Cells
Avril A. Lawshé, Ashley D. Smith, Shalmica R. Jackson, Matthew V. Caple and James S. Ross
Cell Sciences & Development, SAFC Biosciences 2909 Laclede Avenue, Saint Louis, Missouri 63103, USA

Introduction
Undefined components, such as protein hydrolysates, have been routinely used in the A B
biopharmaceutical industry to boost the growth and productivity of recombinant protein- 1.0E+07
EX-CELL™ CD CHO Fusion
Cell Line 1
250 1.0E+07
Cell Line 2

900
EX-CELL™ CD CHO Fusion

producing Chinese Hamster Ovary (CHO) cells in culture. Due to regulatory concerns and 9.0E+06
Competitor A
Competitor B
Competitor C
9.0E+06
Competitor B
Competitor C 800
8.0E+06 200 Competitor D

the variability associated with these components, a need for chemically defined (CD) basal

Average Viable Cells (cells/mL) -

Competitor D 8.0E+06
Competitor E 700

Average Viable Cells (cells/mL) -

Competitor E
EX-CELL™ CD CHO Fusion
7.0E+06 EX-CELL™ CD CHO Fusion 7.0E+06 Competitor B

formulations has emerged. SAFC Biosciences (SAFC) has offered CD formulations in the 600
Competitor A

rIgG (mg/L) - bars

rIgG (mg/L) - bars

Competitor B Competitor C
6.0E+06 150 6.0E+06
Competitor C Competitor D
500

lines
Competitor D Competitor E

past and has now expanded on that knowledge and expertise to develop a novel chemically 5.0E+06 5.0E+06

lines
Competitor E
400
4.0E+06 100 4.0E+06

defined formulation that yields both good growth and productivity for cultured CHO cells. 3.0E+06 3.0E+06
300

This most recent medium, EX-CELL™ CD CHO Fusion, has shown enhanced performance 2.0E+06

1.0E+06
50 2.0E+06
200

100
1.0E+06

in batch culture alongside legacy SAFC formulations as well as competitive performance 0.0E+00 0 0.0E+00 0
0 1 2 3 4 5 6 7 8 9 0 1 2 3 4 5 6 7 8 9 10
against formulations available from external vendors. In addition, EX-CELL™ CD CHO Fusion Days Days

shows good performance in fed-batch culture utilizing various plant hydrolysates as well as
proprietary chemically defined feeds. C
Cell Line 3
8.0E+06 800
EX-CELL™ CD CHO Fusion
Competitor A

Materials and Methods 7.0E+06 Competitor B

Competitor C
700

Average Viable Cells (cells/mL) -

Competitor D
6.0E+06 Competitor E
600

Cell lines and media: EX-CELL™ CD CHO Fusion

rIgG (mg/L) - bars

5.0E+06 Competitor A 500
Competitor B

lines
Competitor C

Four recombinant IgG-producing test cell lines (Table 1) were initially maintained in 4.0E+06 Competitor D
Competitor E
400

EX-CELL™ CD CHO Fusion (SAFC Biosciences) and supplemented appropriately. 3.0E+06 300

2.0E+06 200

CELL LINE PARENTAL MEDIA SUPPLEMENT 1.0E+06 100

0.0E+00 0
1 CHO-S Derived 6mM L-Glutamine 0 1 2 3 4 5 6 7 8 9 10 11
Days

2 CHO-S Derived 6mM L-Glutamine

3 CHOK1SV* 25µM methionine sulphoximine Figure 2. EX-CELL™ CD CHO Fusion compared to five competitor formulations.

4 CHOK1SV* 25µM methionine sulphoximine A) Cell Line 1, B) Cell Line 2, C) Cell Line 3. Results show the differences in the viable cell densities and cumulative
IgG productions of each cell line. In each experiment, EX-CELL™ CD CHO Fusion is competitive for cell growth and
*Licensed from Lonza Biologics is superior for productivity. The overall performance of EX-CELL™ CD CHO Fusion demonstrates its versatility among
cell lines and competitiveness with other formulations available for CHO cell culture. Note: Competitor A did not
survive the adaptation process for Cell Line 2 so was not included in the assay.
All legacy SAFC Biosciences formulations compared to EX-CELL™ CD CHO Fusion
(SAFC Biosciences) were chemically defined. A B
Cell Line 1 Cell Line 2

For competitor assays, five CD formulations were tested alongside EX-CELL™ CD CHO 7.0E+06
Glucose only
250 8.0E+06
Glucose only
800

Soy
Fusion (SAFC Biosciences). All stocks were adapted over at least five passages in each test 6.0E+06 Yeast
Wheat
7.0E+06
Soy
Yeast
Wheat
700
Average Viable Cells (cells/mL) - lines

Average Viable Cells (cells/mL) - lines

200
condition before initiation of growth and productivity assays. 5.0E+06
Feed 1
Feed 2
Feed 3
6.0E+06
Feed 1
Feed 2
Feed 3
600

Glucose only Glucose only

rIgG (mg/L) - bars

rIgG (mg/L) - bars

Soy 5.0E+06 Soy 500
150
In the fed-batch assays, EX-CELL™ CD CHO Fusion (SAFC Biosciences) was supplemented 4.0E+06 Yeast
Wheat
Yeast
Wheat
Feed 1 4.0E+06 Feed 1 400
on days two and four of culture with 3 g/L glucose and either 10% initial culture volume of 3.0E+06
Feed 2
Feed 3 100
Feed 2
Feed 3
3.0E+06 300
proprietary chemically defined feed or 1g/L hydrolysates, where appropriate. 2.0E+06
2.0E+06 200
50

For each assay, cultures were inoculated directly from stocks into TPP® (Techno Plastic 1.0E+06
1.0E+06 100

Products AG) 50ml bioreactor tubes and conditions were averaged after being evaluated in 0.0E+00
0 1 2 3 4 5 6 7 8
0 0.0E+00 0
0 1 2 3 4 5 6 7 8 9 10
duplicate. Cultures were seeded at 2-3 E5 cells per milliliter and terminated when viabilities Days Days

decreased to less than 70%.

C D
Productivity assay: Cell Line 3 Cell Line 4

6.0E+06 900 6.0E+06 1800

Human IgG: IgG concentrations were quantitated using the Octet QK Bio-Layer Interferometer Glucose only
Soy 800
Glucose only
Soy 1600
Yeast
(ForteBio). 5.0E+06 Wheat 5.0E+06
Yeast
Wheat
Average Viable Cells (cells/mL) - lines
Average Viable Cells (cells/mL) - lines

Feed 1 700 Feed 1 1400

Feed 2 Feed 2
Feed 3 Feed 3
4.0E+06 Glucose only 600 4.0E+06 1200
Glucose only

rIgG (mg/L) - bars

rIgG (mg/L) - bars

Soy Soy
Yeast Yeast
Results 3.0E+06
Wheat
Feed 1
500
3.0E+06
Wheat
Feed 1
1000

Feed 2 400 Feed 2 800

Feed 3 Feed 3

A Cell Line 1
2.0E+06 300 2.0E+06 600

8.0E+06 300
200 400
14360C
Average Viable Cells (cells/mL) -

1.0E+06 1.0E+06
7.0E+06 C4726
250 100 200
C1490
6.0E+06
rIgG (mg/L) - bars

EX-CELL™ CD CHO Fusion 0.0E+00 0 0.0E+00 0

14360C 200 0 1 2 3 4 5 6 7 8 9 10
5.0E+06 0 1 2 3 4 5 6 7 8 9 10
C4726 Days Days
lines

4.0E+06 C1490 150

EX-CELL™ CD CHO Fusion
3.0E+06
100 Figure 3. Fed-batch comparison in EX-CELL™ CD CHO Fusion using three plant hydrolysates and three
2.0E+06 chemically defined feeds.
50
1.0E+06
A & B) Cell Lines 1 and 2, C & D) Cell Lines 3 and 4. While the degree to which feeds improve culture performance
0.0E+00 0
0 1 2 3 4 5 6 7 8 9 10 is cell line dependent, the data show that EX-CELL™ CD CHO Fusion can be used in combination with multiple
Days defined and undefined feeds to enhance growth and productivity of CHO cells.
B Cell Line 2
5.0E+06 1000
14360C
900
Conclusions
Average Viable Cells (cells/mL) -

C4726
4.0E+06 C1490 800
• SAFC Biosciences has developed a novel chemically defined formulation called EX-CELL™
rIgG (mg/L) - bars

EX-CELL™ CD CHO Fusion

700
14360C
3.0E+06 C4726 600
CD CHO Fusion.
C1490 500
lines

2.0E+06
EX-CELL™ CD CHO Fusion
400 • EX-CELL™ CD CHO Fusion exceeds the overall performance of legacy SAFC chemically
300
defined catalog offerings.
1.0E+06 200
100 • When used alongside chemically defined competitor formulations, EX-CELL™ CD CHO
0.0E+00 0
0 1 2 3 4 5 6 7 8 9 10 Fusion achieves competitive growth and superior cumulative productivity.
Days
• EX-CELL™ CD CHO Fusion can be used as a robust platform formulation that, when
Figure 1. EX-CELL™ CD CHO Fusion compared to legacy SAFC Catalog formulations. complimented with the appropriate CD feed, effectively replaces the need for undefined
A) Cell Line 1, B) Cell Line 2. CHO cells were screened with all the chemically defined catalog formulations currently components to yield increased IgG production.
available from SAFC Biosciences. As seen from improvements in both growth and productivity, EX-CELL™ CD CHO
Fusion demonstrates the progress that has been made in the development of chemically defined formulations that
can produce high cell densities and IgG titers.

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