In vitro targeted killing of human endothelial cells by co-incubation of human serum and NGR peptide conjugated human albumin protein bearing alpha (1-3) galactose epitopes.

The NGR/alpha1,3Gal-HSA peptide was designed to specifically target CD13 positive cells and induce cell lysis. NGR is the targeting component of the peptide in that it binds the CD13 isoform (aminopeptidase) that is expressed in tumor vessels. Galactose alpha1,3-galactose terminal carbohydrate epitope (alpha1,3Gal) induces a strong antibody reaction in human and Old World Monkeys and in vivo, this reaction leads to organ rejection. The human serum albumin (HSA) bearing alpha1,3Gal epitope was therefore used to lyse cells. In the present study, we were able to demonstrate that NGR/alpha1,3Gal-HSA binds CD13 positive human umbilical vein endothelial cells (HUVEC). We also found by live/dead fluorescent staining that NGR/alpha1,3Gal-HSA was able to induce lysis of HUVECs upon incubation with human serum. Therefore, by conjugating NGR to HSA bearing alpha1,3Gal epitopes, we are able to specifically target and lyse cells expressing CD13. This strategy may be potentially useful in tumor anti-angiogenesis therapy.

Bcl-2 targeting siRNA expressed by a T7 vector system inhibits human tumor cell growth in vitro.

A T7 promoter driven siRNA expression vector system (Bcl-2/T7) that targets Bcl-2 mRNA in MCF-7 human cancer cells was designed in the present study. In the presence of prebound T7 RNA polymerase, successful expression of Bcl-2 siRNA as well as its function was demonstrated via cell proliferation assays, Bcl-2 Elisa, and TUNEL assay. MCF-7 breast cancer cells transfected with Bcl-2/T7 show decreased levels of Bcl-2 expression at the protein level as well as decreased cell proliferation. Also, the number of apoptotic cells was increased in cells expressing Bcl-2 siRNA. Previous studies have shown that Bcl-2 levels are increased in a large number of different types of cancer. Therefore, the ability of Bcl-2/T7 to produce functional Bcl-2 siRNA in breast cancer cells suggests a potential role for this delivery system in cancer gene therapy.