Purification of clinical-grade recombinant HSP65-MUCI fusion protein.
HSP65-MUCI is a fusion protein between BCG (Bacille Calmette-Guerin)-derived HSP65 (heat-shock protein 65) and human MUCI (mucin I) VNTR (variable number of tandem repeats)-domain peptides that has shown antitumour efficacy. China's Food and Drug Administration has recently approved a Phase I clinical trial using HSP65-MUCI for the treatment of MUCI-positive breast cancer. In order to produce sufficient quantities of clinical-grade HSP65-MUCI, we established a pilot-scale purification scheme comprising two steps of column chromatography: HIC (hydrophobic-interaction chromatography) and IEX (ion-exchange chromatography). The pH values of the buffers used in homogenization and HIC were adjusted to pH 9.0 to maintain protein stability and prevent protein degradation. Using this manufacturing process, we obtained clinical-grade HSP65-MUCI with a yield of 400 mg per 70 g of wet cell pellet and >96% purity.
Cao
Zhao
Z
Department of Molecular Biology, Norman Bethune College of Medicine, Jilin University, Changchun 130021, People's Republic of China.
Feng
Yu
Y
Wei
Hongfei
H
Fang
Mingli
M
Hu
Xiaoping
X
Yu
Yongli
Y
Wang
Liying
L
Wan
Min
M
D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D019496:Cancer Vaccines / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D018834:Chaperonin 60 / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018598:Minisatellite Repeats / Q000235:genetics; D018396:Mucin-1 / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D009163:Mycobacterium bovis; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D019496:Cancer Vaccines; D018834:Chaperonin 60; D018396:Mucin-1; D011993:Recombinant Fusion Proteins; C074119:heat-shock protein 65, Mycobacterium