Combinatorial effects of monoclonal anti-p120 antibody (MAbp120), liposomes and hyperthermia on MCF-7 and LOX tumor cell lines.

The human nucleolar protein p120 is highly expressed in human cancers. Its high expression in breast cancer correlates with a poor prognosis, and its overexpression in 3T3 mouse fibroblasts causes malignant transformation. This study reports that a combination of monoclonal anti-p 120 antibody (MAbp120), liposomes (Lipo), and hyperthermia (HT) resulted in enhanced antitumor effects in cultured human breast adenocarcinoma (MCF-7) and human amelanotic melanoma (LOX) cells. Monoclonal antibody uptake and intracellular localization of the protein p120 were monitored by double labeling indirect immunofluorescence. Cell growth inhibition by the combination of MAbp120 + Lipo + HT was 65% for MCF-7 cells and 96% for LOX cells. When tested on LOX cells, monoclonal antibodies (MAbB23, MAbC23) to other nucleolar proteins (B23, C23) produced only slight cytotoxicity with similar treatment protocols.

Characterization of U6 small nuclear RNA cap-specific antibodies. Identification of gamma-monomethyl-GTP cap structure in 7SK and several other human small RNAs.

The cap structure in human U6 small nuclear (sn)RNA, gamma-monomethylguanosine triphosphate (meGTP), was conjugated to human serum albumin and used as antigen to raise polyclonal antibodies in rabbits. The resulting antibodies reacted specifically with meGTP but not with GTP, GDP, GMP, meGMP, meATP, meCTP, meUTP, or with methyl phosphate in enzyme-linked immunosorbent assay and/or in radioimmunoassays. Although less efficiently, meGDP was also recognized by these antibodies. Indirect immunofluorescence studies with anti-meGTP antibodies showed predominantly nuclear immunofluorescence. Anti-meGTP antibodies immunoprecipitated intact U6 snRNA from a mixture of HeLa cell RNAs. In addition to the U6 snRNA, anti-meGTP antibodies immunoprecipitated several additional small RNAs that varied in length from approximately 50 to 330 nucleotides. These RNAs contained the meGTP cap structure and are structurally distinct from U6 snRNA. One of these meGTP-containing RNAs was found to be previously characterized 7SK RNA; human 7SK RNA synthesized in vitro also contained the same cap structure. Results obtained in this study provide evidence for the presence of gamma-monomethyl-GTP cap structure in a wide spectrum of human cellular RNAs. These antibodies will be useful in studying the structure and function of this new family of small RNAs.