A recombinant human RNASET2 glycoprotein with antitumorigenic and antiangiogenic characteristics: Expression, purification, and characterization

Patricia Smirnoff, Levava Roiz, Boaz Angelkovitch, Betty Schwartz, Oded Shoseyov*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

53 Scopus citations


BACKGROUND. Human RNASET2 is a T2-RNaSe glycoprotein encoded by the RNA-SET2 gene, which is located on chromosome 6 (6q27). Deletion in 6q27 is associated with several human malignancies. METHODS. A synthetic RNASET2 gene that was optimized for expression in the yeast Pichia pastoris was designed according to the cDNA sequence and was cloned under the control of the methanol-induced promoter fused to the α-mating secretion peptide. The recombinant protein was purified from the culture supernatant of transformed P. pastoris through an affinity Sepharose-concanavalin A column. Actin-binding activity was examined by membrane blotting using monoclonal mouse antiactin immunoglobulin M and by cross-linking in solution to G-actin using 1-[3-(dimethylamino)propyl]-3-ethyl-carboimide methiodide. The antiangiogenic activity of RNASET2 (from 0.5 μM to 10 μM) was assessed by a human umbilical vein endothelial (HUVE) cell assay in the presence of 1 μg/mL angiogenin, basic fibroblast growth factor (bFGF), or recombinant human vascular endothelial growth factor (VEGF). Cell colony formation was examined in human colon HT29 cancer cells to assess the antitumorigenic activity of RNA-SET2 or the enzymatic-inactivated RNASET2 (EI-RNASET2) (1 μM each). In an athymic mouse xenograft model, LS174T human cancer cells were injected sub-cutaneously. When tumors were palpable, the mice were treated for 3 weeks with RNASET2 (1 mg/kg), paclitaxel (10 mg/kg or 15 mg/kg), or a combination of the 2 drugs. RESULTS. The recombinant RNASET2 was identified as a 27-kilodalton glycoprotein that possessed the ability to bind actin in vitro. RNASET2 significantly inhibited clonogenicity in HT29 cells. EI-RNASET2 produced a similar effect, suggesting that its antitumorigenic activity is unrelated to its RNase activity. In HUVE cells, RNASET2 inhibited angiogenin-, bFGF-, and VEGF-induced tube formation in a dose-dependent manner. In athymic mice, RNASET2 inhibited the development of an LS174T-derived xenograft by 40%. A synergistic effect was obtained with combined RNASET2 and paclitaxel treatments. CONCLUSIONS. The current results suggested that RNASET2 represents a new class of antitumorigenic and antiangiogenic drugs, and the findings of this study emphasize the advantage of using agents like RNASET2 in combined therapy.

Original languageAmerican English
Pages (from-to)2760-2769
Number of pages10
Issue number12
StatePublished - 15 Dec 2006


  • Actin-binding
  • Antiangiogenic
  • Colon cancer
  • Paclitaxel


Dive into the research topics of 'A recombinant human RNASET2 glycoprotein with antitumorigenic and antiangiogenic characteristics: Expression, purification, and characterization'. Together they form a unique fingerprint.

Cite this