Recombinant endostatin forms amyloid fibrils that bind and are cytotoxic to murine neuroblastoma cells in vitro.
Endostatin is a fragment of collagen XVIII that acts as an endogenous inhibitor of tumor angiogenesis and tumor growth. Anti-tumor effects have been described using both soluble and insoluble recombinant endostatin. However, differences in endostatin structure are likely to cause differences in bioactivity. In the present study we have investigated the structure and cellular effects of insoluble endostatin. We found that insoluble endostatin shows all the hallmarks of amyloid aggregates. Firstly, it binds Congo red and shows the characteristic apple-green birefringe when examined under polarized light. Secondly, electron microscopy shows that endostatin forms short unbranched fibrils. Thirdly, X-ray analysis shows the abundant presence of cross-beta sheets, the tertiary structure that underlies fibrillogenesis. None of these properties was observed when examining soluble endostatin. Soluble endostatin can be triggered to form cross-beta sheets following denaturation, indicating that endostatin is a protein fragment with an inherent propensity to form amyloid deposits. Like beta-amyloid, found in the brains of patients with Alzheimer's disease, amyloid endostatin binds to and is toxic to neuronal cells, whereas soluble endostatin has no effect on cell viability. Our results demonstrate a previously unrecognized functional difference between soluble and insoluble endostatin, only the latter acting as a cytotoxic amyloid substance.
Kranenburg
Onno
O
Department of Medical Oncology, University Medical Center Utrecht, Heidelberglaan 100, 3584 CX Utrecht, The Netherlands.
Kroon-Batenburg
Loes M J
LM
Reijerkerk
Arie
A
Wu
Ya-Ping
YP
Voest
Emile E
EE
Gebbink
Martijn F B G
MF
D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry; D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology; D017209:Apoptosis; D003094:Collagen / Q000737:chemistry / Q000494:pharmacology*; D043170:Collagen Type XVIII; D003603:Cytotoxins / Q000737:chemistry / Q000494:pharmacology*; D043169:Endostatins; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000187:drug effects*; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D012995:Solubility; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
D000682:Amyloid; D000970:Antineoplastic Agents; D043170:Collagen Type XVIII; D003603:Cytotoxins; D043169:Endostatins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D003094:Collagen