A MONOCLONAL-ANTIBODY WITH HIGH-AFFINITY FOR A NEO-ANTIGENIC SITE IN FIBRINOGEN DEGRADED BY POLYMORPHONUCLEAR LEUKOCYTE-DERIVED ELASTASE

Elastase, released by stimulated polymorphonuclear leukocytes (PMN), i s thought to play an important role in the pathogenesis of chronic obs tructive pulmonary disease (COPD) especially pulmonary emphysema. A te st that can detect release of elastase activity from PMN would be valu able to monitor therapy or to identify patients at risk. The authors a imed to isolate and characterize monoclonal antibodies (mAb) with a hi gh affinity for a neo-antigenic determinant in a high-molecular weight degradation product of fibrinogen (Fbg) generated by PMN-derived elas tase. Using synthetic peptides, they mimicked the new amino or carboxy terminal sequences of the A alpha-, B beta- and gamma-chains of Fbg t hat are generated by elastase. These synthetic peptides (A alpha 22-36 , A alpha 350-360, B beta 44-55, and gamma 295-305), uni-directionally coupled to a carrier protein, were used to generate mAb specific for elastase-degraded fibrinogen (EDF). mAb that appeared to be specific f or a neo-antigenic determinant (neotope) consisting of the new amino t erminal amino acid(s) of the Fbg A alpha-chain that is generated by el astase activity were isolated only with the A alpha 22-36 synthetic pe ptide. One mAb, designated as EF1-4, was further characterized and had an approximately 75-fold higher affinity for EDF, as compared with Fb g, in solution. Using the other peptides, no mAb specific for elastase generated fibrinogen degradation products were obtained. Incubation o f immobilized fibrin(ogen) with stimulated PMN resulted in the generat ion of this specific neotope recognized by mAb EF1-4; the protease inh ibitors alpha(1)-proteinase inhibitor (alpha(1)-PI) and antileukoprote ase (ALP) decreased the generation by PMN of this neotope on Fbg. Fibr inogen degradation products generated by plasmin or by other leukocyte proteases, i.e. cathepsin G and proteinase 3, did not react with mAb EF1-4.