Enabling low cost biopharmaceuticals: high level interferon alpha-2b production in Trichoderma reesei.
BACKGROUND
The filamentous fungus Trichoderma reesei has tremendous capability to secrete over 100 g/L of proteins and therefore it would make an excellent host system for production of high levels of therapeutic proteins at low cost. We have developed T. reesei strains suitable for production of therapeutic proteins by reducing the secreted protease activity. Protease activity has been the major hindrance to achieving high production levels. We have constructed a series of interferon alpha-2b (IFNα-2b) production strains with 9 protease deletions to gain knowledge for further strain development.


RESULTS
We have identified two protease deletions that dramatically improved the production levels. Deletion of the subtilisin protease slp7 and the metalloprotease amp2 has enabled production levels of IFNα-2b up to 2.1 and 2.4 g/L, respectively. With addition of soybean trypsin protease inhibitor the level of production improved to 4.5 g/L, with an additional 1.8 g/L still bound to the secretion carrier protein.


CONCLUSIONS
High levels of IFNα-2b were produced using T. reesei strains with reduced protease secretion. Further strain development can be done to improve the production system by reducing protease activity and improving carrier protein cleavage.
VTT Technical Research Centre of Finland Ltd., Espoo, Finland. christopher.landowski@vtt.fi.
Christopher P
CP
Landowski
VTT Technical Research Centre of Finland Ltd., Espoo, Finland.
Eero
E
Mustalahti
Novartis Pharma AG, Basel, Switzerland.
Ramon
R
Wahl
Novartis Pharma AG, Basel, Switzerland.
Laurence
L
Croute
VTT Technical Research Centre of Finland Ltd., Espoo, Finland.
Dhinakaran
D
Sivasiddarthan
VTT Technical Research Centre of Finland Ltd., Espoo, Finland.
Ann
A
Westerholm-Parvinen
Novartis Pharma AG, Basel, Switzerland.
Benjamin
B
Sommer
Novartis Pharma AG, Basel, Switzerland.
Christian
C
Ostermeier
Novartis Pharma AG, Basel, Switzerland.
Bernhard
B
Helk
Glykos Finland Oy, Helsinki, Finland.
Juhani
J
Saarinen
VTT Technical Research Centre of Finland Ltd., Espoo, Finland.
Markku
M
Saloheimo
D019149:Bioreactors; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000096:biosynthesis* / Q000191:economics / Q000235:genetics; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000191:economics / Q000235:genetics; D014242:Trichoderma / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000378:metabolism
D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D014361:Trypsin Inhibitors; D010447:Peptide Hydrolases
Cytokines; Filamentous fungi; Interferon; Proteases; Therapeutic proteins; Trichoderma reesei