© 1998 by Oxford University Press
Journal Of The National Cancer Institute, Vol 90, 1787-1791, Copyright © 1998 by Oxford University Press
DL Newton and SM Rybak
BACKGROUND: Preparations of human chorionic gonadotropin (hCG) have been
shown to exhibit anti-Kaposi's sarcoma (KS) activity, but the identity of
the responsible agent(s) remains controversial. One candidate agent is an
eosinophil-derived neurotoxin (EDN)-like polypeptide that contaminates
preparations of hCG. We have genetically engineered a unique form of hEDN,
which is a ribonuclease, and have evaluated the cytotoxic effects of the
recombinant protein on KS Y-1 cells and on cells of other cancer types.
METHODS: The amino-terminus of hEDN was extended by four amino acid
residues, corresponding to the proximal part of the hEDN signal peptide
(serine, leucine, histidine, and valine; positions -4 to -1, respectively),
by use of the polymerase chain reaction and an hEDN complementary DNA. The
recombinant protein was isolated from bacterial inclusion bodies. The
cytotoxic activity of this hEDN variant, (-4)rhEDN, was tested on KS Y-1
cells and human glioma, melanoma, breast carcinoma, and renal carcinoma
cells. RESULTS: Approximately half of the anti-KS activity in a crude
commercial preparation of hCG was associated with a polypeptide that
reacted with anti-recombinant-hEDN (rhEDN) polyclonal antibodies. Although
rhEDN protein displayed little cytotoxicity against KS Y-1 cells (IC50 [50%
inhibition concentration] = >100 microg/mL), (-4)rhEDN markedly
inhibited cell viability (IC50 = 6 microg/mL). Neither version of rhEDN
inhibited the viability of other tested human cancer cell types.
CONCLUSIONS: A four amino acid extension of the amino-terminus of rhEDN
confers cytotoxicity against KS Y-1 cells in vitro. Design of the (-
4)rhEDN variant was based on the sequence of a natural human protein
associated with hCG. Our results suggest that (-4)rhEDN is one of the
agents in hCG responsible for anti-KS activity. A purified molecule is thus
available for in vitro and in vivo mechanistic and, possibly, future
clinical studies.
ARTICLES
Unique recombinant human ribonuclease and inhibition of Kaposi's sarcoma cell growth
Intramural Research Support Program, SAIC Frederick, National Cancer Institute-Frederick Cancer Research and Development Center, MD 21702- 1201, USA.
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