Enhanced reactive lysis of paroxysmal nocturnal hemoglobinuria erythrocytes. Studies on C9 binding and incorporation into high molecular weight complexes.

Abstract

The human disease paroxysmal nocturnal hemoglobinuria (PNH)' may be regarded as an experiment of nature that has already provided, and should continue to provide, unique insights on the cytolytic function of complement (C) and its regulation (1-4). The studies of our laboratory have been focused particularly on elucidating the process by which the most sensitive type of PNH erythrocyte (E) (designated type III) are abnormally sensitive to in vitro reactive lysis, i.e., to membrane attack initiated by purified C5b6, C7, C8, and C9 in the absence of the earlier activation and amplification steps (5).In an earlier report (6), we presented evidence that for a given input of purified C5b6, type III PNH E (PNH-III E) did not bind '21 I-C7 to a greater extent than normal human E. Rather, the data indicated that PNH E underwent two-fourfold greater lysis for a given number of C7 molecules bound. Compatible results were obtained in less extensive, unpublished studies with radiolabeled C5b6. Compelling independent evidence from other laboratories (7) indicates that the basic unit of the membrane attack complex involves one molecule each of C5b, C6, C7, and C8 in the formation of the C5b-8 complex, to which a variable number of C9 molecules then bind to produce the fully effective membrane attack complexes. Therefore, although C8 uptake has not been directly measured in our system, it seems justified to infer that in reactive lysis the assembly of C5b-8 complexes on PNH-III E from the fluid phase is quantitatively normal . The excessive lysis of the PNH E by the full C5b-9 sequence could result from a quantitative abnormality affecting C9 binding and/or polymerization within the C5b-9 complex, or from a qualitative difference in the disposition or function of the terminal C components after they have become bound to the PNH membrane. The previously reported deficiency of a membrane complement regula-

Paroxysmal nocturnal hemoglobinuria type III. Lack of an erythrocyte membrane protein restricting the lysis by C5b-9.

“…The reason for the enhanced susceptibility is still under investigation; meanwhile, enhanced binding ofthe components ofthe attack phase has been suggested (35). Other authors, however, reported that in absence of C3 (i.e., using isolated C5b6, C7, C8 and C9) the uptake and incorporation ofthe components into the membrane-attack complex is equal to that ofnormal erythrocytes (36). When, however, whole serum was used, enhanced C9 uptake was seen as well as enhanced C9 polymerization (20).…”

“…Whether this band represents another form ofC8bp or a split product is not yet clear; however, it most likely indicates a form ofC8bp, probably identical with inhibitory protein described by Zalman and Muller-Eberhard (36).…”

Erythropoietin treatment during complement inhibition with eculizumab in a patient with paroxysmal nocturnal hemoglobinuria

“…[2][3][4][5] Currently, no approved therapies effectively reduce intravascular hemolysis in PNH, and patients are often dependent on transfusions to artificially support hemoglobin levels. 2 Eculizumab is a novel humanized monoclonal antibody directed against the complement protein C5.…”