DEVELOPMENT OF A RADIOIMMUNOASSAY FOR A PSEUDONONAPEPTIDE BOMBESIN GRP ANTAGONIST WITH ANTITUMOR-ACTIVITY

Bombesin-like and GRP-like peptides may act as autocrine growth factor s in the proliferation of some cancers. A pseudononapeptide bombesin a ntagonist, [D-Tpi(6),Leu(13) Psi(CH2NH)-Leu(14)]bombesin(6-14), and re lated analogs synthesized in our laboratory significantly inhibit tumo r growth in various cancer models. A radioimmunoassay (RIA), suitable for determination of RC-3095 and its congeners in unextracted serum, w as developed in order to facilitate further experimental and clinical evaluation of this bombesin/GRP receptor antagonist for the treatment of various tumors. Antibodies were generated against RC-3095 and Des-T pi(1)-RC-3095, conjugated to bovine serum albumin with glutaraldehyde. Antiserum JH-631b was selected for further experiments based on the a ntibody characterization. At an antiserum dilution of 1:189000, this a ntibody bound approximately 50% of 7 fmol of added radiolabeled Tyr(1) -RC-3095. The antibody crossreacted with C-terminal fragments of RC-30 95. Fragments without the C-terminus and naturally existing peptides o f the bombesin family or structurally unrelated peptides did not cross -react. The minimum detectable dose of RC-3095 was 0.4 pg/tube. Intra- and interassay coefficients of variation ranged from 3.2 to 4.4% and from 5.6 to 12.8%, respectively. The RIA is suitable for direct determ ination of RC-3095 in serum. The RIA should be of value for monitoring levels of this analog in serum during long-term therapy. (C) Munksgaa rd 1995.