No abstract

SUMMARYWe recently described the identity of Ihe small cell lung cancer (SCI.C) cluster-w4 antigen and the human B cell ditlcrentiation marker CD24, a glycosylphosphatidylinosilol (GPI)-anchored, highly glycosylalcd surface molecule ofonly 31 -35 amino acids [15]. The specificities of three anti-cluster-w4 and of eleven anti-CD24 MoAbs have been investigated with respect to their binding capacity lo the protein core of cluster-w4/CD24 antigen. Four overlapping peptides spanning ihis protein core were synthesized. MoAbs shown to bind to two overlapping peptides by antibody binding inhibition using the cluster-w4/CD24-positive SCLC cell line SW2 and by direct peptide binding detecied in an ELISA were investigated in more detail. To determine the exact epitopes recognized by these MoAbs. an epitope mapping assay using peptides synthesized onto polyethylene pins was established. The three anti-cluster-w4 MoAbs SWA 11, SWA2I and SWA22 and the anti-CD24 MoAbs OKB2 and ALB9 recognized the same short leucine-alanine-proline (LAP) sequence in an area without potential glycosytation sites close lo the GPI anchor oi the protein core of the cluster-w4/CD24 antigen.Keywords epitope mapping monoclonal aniibody SWA 11 clusler-w4 antigen CD24 antigen small cell lung cancer

We have previously developed 3 monoclonal anti-idiotypic antibodies (Ab2) of LOU rat origin directed against the binding site of the murine monoclonal IgM LAM8, which recognizes the small-cell lung carcinoma (SCLC)-associated sialoglycoprotein antigen sGP 90-135. The aim of this study was to compare the efficiencies of these 3 Ab2, designated LY8-229, LX8-531 and LX8-632, to induce antigen-specific immunity in different animal species without prior exposure of the recipients to the nominal antigen, and thereby possibly select an Ab2 candidate for active immunotherapy against SCLC in patients. The feasibility of this approach was further evaluated by a serological analysis of patients with SCLC compared with healthy individuals, in whom the spontaneous antibody reactivities against SCLC cell lines and Ab2 were tested. LY8-229 was shown to be the most effective Ab2 in inducing antigen-specific antibodies in BALB/c mice, CBA/J/Zur mice and one NZW rabbit. Furthermore, LY8-229 was the only Ab2 against which significantly elevated idiotype-specific antibody reactivities existed in sera of patients with SCLC. These reactivities correlated positively with binding to antigen-positive tumor cells. Our findings suggest that LY8-229 represents in its reactivity pattern the nominal SCLC antigen in humans also, and therefore may be of diagnostic and possibly therapeutic relevance for patients with SCLC.

Lehmann H-P. New concepts in systemic autoimmunity testing. Scand J Clin Lab Invest 2001;61(Suppl 235):84-90.Diagnosis of systemic autoimmune diseases is highly complex, and it is becoming increasingly difficult to make assumptions about the functional roles and diagnostic significance of autoantibodies. The latter is mainly due to the fact that results from different assay systems are not interchangeable. A laboratory "gold standard" which helps the clinician to differentiate irrelevant autoimmune phenomena from significant autoimmune diseases at an early stage, is clearly missed. To meet this challenge, a rheuma entrance screening (RES) assay toolbox is proposed based on fully-automated enzyme immunoassay (EIA) technology on one system for the clinical and routine laboratory. The RES concept is intended to cover the most important syndromes of systemic rheumatic diseases, i.e. collagenosis, early rheumatoid arthritis, early osteoarthritis, anti-phospholipid syndrome and inflammation. The serological part of diagnosis of these diseases comprises testing for anti-nuclear antibodies (ANA), rheumatoid factor (RF), low levels of C-reactive protein (CRP), and disease-specific anti-phospholipid antibodies, e.g. anti-beta-2 glycoprotein I (anti-2 GPI). To eliminate the known problems of varying assay systems in this field, a novel, objective, rapid and reproducible approach to screen for such analytes in patient serum or plasma more efficiently is the application of EIAs on the fully-automated immunoassay analyser COBAS Ò CORE (Roche Diagnostics GmbH, Mannheim, Germany). The combined use of the RES (COBAS Ò CORE HEp2 ANA EIA, COBAS Ò CORE RF EIA Quant, COBAS Ò CORE CRP EIA Quant and COBAS Ò CORE Anti-2 GPI EIA) is intended for patients sent to the laboratory with the primary suspicion of harbouring a systemic rheumatic disease.

The therapeutic efficacy of an immunotoxin, SWAII-SPDB-dg.ricin A chain, recognizing the leukocyte-differentiation antigen CD24, was evaluated against SCLC cell lines in tissue culture and in 2 nude-mouse models. The first model used conventional s.c. solid-tumor xenografts. The second used small tumor-cell deposits established in s.c. implanted sponge matrices and allowed us to directly estimate the killing efficiency of the immunotoxin under experimentally defined conditions in vivo. It also mimics the clinical setting of disseminated tumor cells which form the basis of residual disease in SCLC. The cytotoxic potency of SWAII-SPDB-dg.ricin A chain was demonstrated in tissue culture by the inhibition of 3H-leucine incorporation and by the selective elimination of CD24-positive tumor cells in clonogenic assays. In nude mice, SWAII-SPDB-dg.ricin A chain was cleared from the blood circulation with biphasic kinetics: an initial alpha phase of 1 hr and a second beta phase of 20.5 hr. Following i.v. injection of a dose equivalent to 30% of the LD50, the immunotoxin delayed the growth of SW2 solid-tumor xenografts by 16 days. The therapeutic efficacy of SWAII-SPDB-dg.ricin A chain was further demonstrated by the selective elimination of clonogenic SW2 cells from small tumor-cell deposits established in sponge matrices. Regrowth of the solid tumors after the initial response and the clonogenic activity in the sponge-derived cell population were mediated by CD24-positive cells, excluding the selection of CD24-negative mutants during immunotoxin therapy.