PEGylated recombinant human tumor necrosis factor alpha: preparation and anti-tumor potency.
OBJECTIVE
To assess the merits of polyethylene glycol-modified recombinant human tumor necrosis factor alpha (PEG-rHuTNF-alpha).


METHODS
The rHuTNF-alpha was modified with N-succinimidyl succinate monomethoxy polyethylene glycol (SS-PEG) of three different molecular weights. The PEG-rHuTNF-alpha was separated into fractions of various molecular weights by gel filtration chromatography. In vitro activities of various fractions were determined with L929 cell assay and in vivo anti-tumor potencies of main fractions were studied with respect to necrosis of S-180 solid tumor.


RESULTS
The rHuTNF-alpha could be modified using SS-PEG under mild conditions. The main fraction of PEG5000-rHuTNF-alpha contained four PEG molecules, and PEG12000-rHuTNF-alpha and PEG20000-rHuTNF-alpha contained two PEG molecules, respectively. There was a higher activity when rHuTNF-alpha was coupled to less numbers of the same molecular weight PEG molecules. When PEG-rHuTNF-alpha was of the same molecular weight, rHuTNF-alpha modified with bigger molecular weight PEG molecules had a higher activity. PEG-rHuTNF-alpha was resistant to proteolysis, and over 70 % activity remained after 8 h, but the activity of rHuTNF-alpha was time-dependently diminished by incubation with bovine trypsin. PEG5000-rHuTNF-alpha (1500 IU per mouse) had a similar anti-tumor potency compared with rHuTNF-alpha (3000 IU per mouse). PEG12000-rHuT NF-alpha (1500 IU per mouse) had an increased anti-tumor potency compared with rHuTNF-alpha (3000 IU per mouse). In particular, PEG20000-rHuTNF-alpha at a dose of 1500 IU per mouse had a higher anti-tumor potency than rHuTNF-alpha at a dose of 6000 IU per mouse.


CONCLUSIONS
PEG-modified rHuTNF-alpha could be more suitable for therapeutic use
Li
Y P
YP
Department of Pharmaceutics, School of Pharmacy, Fudan University, Shanghai 200032, China.
Pei
Y Y
YY
Ding
J
J
Shen
Z M
ZM
Zhang
X Y
XY
Gu
Z H
ZH
Zhou
J J
JJ
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D008297:Male; D051379:Mice; D011092:Polyethylene Glycols; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000494:pharmacology; D012510:Sarcoma 180 / Q000188:drug therapy / Q000473:pathology; D013388:Succinimides; D014407:Tumor Cells, Cultured / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry / Q000494:pharmacology*
D000970:Antineoplastic Agents; D011994:Recombinant Proteins; D013388:Succinimides; D014409:Tumor Necrosis Factor-alpha; D011092:Polyethylene Glycols; C069871:methoxypolyethylene glycol succinimidylsuccinate