Posttranslational processing of endogenous and of baculovirus-expressed human gastrin-releasing peptide precursor.

The 27-amino-acid gastrin-releasing peptide (GRP1-27) is a neuropeptide and growth factor that is synthesized by various neural and neuroendocrine cells. The major pro-GRP hormone (isoform I) contains both GRP1-27 and a novel C-terminal extension peptide termed pro-GRP31-125. In order to define potentially active neuropeptides that could be generated from this novel protein domain, we analyzed the posttranslational processing of endogenous human pro-GRP1-125 in a small-cell lung cancer cell line. Because such studies are much easier in an overexpression system, we investigated at the same time the posttranslational processing of baculovirus-expressed human pro-GRP1-125 in an insect ovary cell line. In the small-cell lung cancer cell line, GRP1-27 was cleaved as expected from the endogenous prohormone at a pair of basic amino acids (29 and 30) and alpha-amidated at its C-terminal methionine; however, a number of novel peptides were generated by additional cleavages in the pro-GRP31-125 domain. In the insect ovary cell line, GRP1-27 was cleaved from the expressed prohormone by a different mechanism, as were a number of other peptides that appeared to be similar in size to those produced by the human neuroendocrine tumor cell line. These data show for the first time that an insect ovary cell line that is widely used to overexpress proteins can process a human neuropeptide precursor. They also reveal the existence of novel pro-GRP-derived peptides that are candidates for biologically active ligands

Monoclonal antibodies for the in vitro detection of small cell lung cancer metastases in human bone marrow.

Three rat monoclonal antibodies were selected for the immunodetection of small cell lung cancer metastases in bone marrow and other hematologic samples. By membrane immunofluorescence, they define three distinct surface antigens here termed lung cancer-associated antigens or LCAs. The latter are widely expressed on small cell lung cancer and non-small cell lung cancer cells/cell lines, but not detectable on a variety of normal and transformed bone marrow, blood and lymphoid cells. Anti-LCA1 (IgM) is similar to the many anti-lacto-N-fucopentaose III IgM antibodies rasied against human tumors. In contrast, anti-LCA2 (IgG2b) and anti-LCA3 (IgG2a) define surface proteins of 29, 32, 41 and 98 kilodaltons, respectively, that have not been reported earlier. These three reagents have immunodiagnostic potential, since in combination they label all 49 lung cancer cell lines tested. Their ability to detect lung cancer metastases in patient's bone marrow samples is documented in an accompanying paper