Complete primary sequences of three λ immunoglobulin light chains in myelomas with light chain deposition disease

328

Immunotherapy

24 June 1997 - Poster presentations

of allergy

The preliminary examination consisted of estimation of the clinical status and cytomorphological, histological. cytogenetic analyses. The final diagnosis has been made taking into account the detailed analysis of the results according to the Common European-American Classification of lymphoproliferations (1QQ5). 9 persons (mean age 39, 55% men) have been examined prior to the treatment. All of them had absolute lymphocytosis over 5 x IO9 I; strengthening in a lymphoid sprout of the bone mamw was over 25%. 6 patients had B-cell proliferation, 3 of them had B-cell lymphoma, 5 persons - B-cell chronic iympholeukosis. The peculiarities of the immunological phenotype were as follows: HLA-DR, CD5, CD20, CD22. T-cell proliferation was diagnosed in one patient: lymphoma resulted in leukoma. The immunological phenotype was CD3, CD5, CD4, CD25, CD36. Determination of the immunological phenotype of the blood cell elements allowed to define a tumour of the immune system. The above said allowed to carry out the cytostatic therapy according to a proper diagnosis.

1P.5.12.20 1 Complete primary sequences of three X lmmunoglobulin light chains in myelomas with light cheln deposition disease C. Decourt’, G. Touchard2, J.L. Preud’homme*, H. Beaufils3. M.-C. Diemert3, M. Cogn6 ‘. lLaboratoire d’/mmuno/ogie, CNRS EP7 IS, Faculaede M&ecine et C.H.R. U. Dupuytren, F87025 Limoges, France, 3LaLnxatoire d’lmmuncchimie, Laboratoire d’Anatomie Pathologique et Service de NBphmiogie, C.H.U. La Piti&Salp&ri~re, France, * Lsbotatoire d’lmmundcgie, Labomtoire d’Anatomie Pathologique ef Service de NBphrologie, C.H. U. La Miletrie, poitiers, Fence Non-amyloid monoclonal immunoglobulin deposition diseases (MIDD) are severe complications of plasma cell dyscrasias, characterized by deposition in various organs of amorphous monoclonal immunoglobulin-related material, mostly light chain (LC), then termed LC deposition disease (LCDD). By contrast to AL-amyloidosis, LCDD is as yet pooffy documented and only a few sequences of LCs involved in LCDDs. all of the K tvoe, have been so far reoorted. We herein report the first prim&y sequences’oi monoclonal LCs responsible for non-amyloid A light chain deposits in three cases which may somewhat differ from the typical ( K LCDD and lack the usual prominent glomerulosclemsis pattern featuring non amyloid K deposits. The sequences were deduced from those of the corresponding complementary (c) DNAs in the bone marmw plasma cells. For each chain, products of three independent amplifications by polymerase chain reaction (PCR) were sequenced and found to be identical. For the first two cases, BOU and RX, the A mRNAs encoding the LCs showed a normal structure consisting of VA2 segments, not deriving from the same germline genes, rearranged to JAXAP. Both chains presented several unusual features and BOUshowed a four amino acids’ deletion within the CDRl region. For the third case, LOF:monoclonal A LC deposits were found associated with complement in the patient’s kidney, the LC consisted in a VA3 segment rearranged to JA2CA2. Interestingly, this VA3 was encoded by the same germline gene as another monoclonal A LC (AUS) that we previously found involved in an unusual form of extra-membranous AL-amyloidosis deposits. Although these two chains weren’t implicated in the same pathology, it is worth noting that the deposits had similar locations and that the proteins shared some structural peculiarities, which had never been reported before for any other A LC. These primary sequences are the first step of such a study and exact comprehension of the mechanisms by which LCs can form deposits will await for molecular modelisation in 3D of the proteins.

non-responsive. Mast cell activation triggered by FaRI clustering has been intensively studied, while the action modes of the anaphyiatoxic psptides is being resolved only recently and studied as a process independent of IgE-mediated triggering. Since these stimuli might be present simultaneously in vivo, we investigated the effect of C3a and C5a peptides on IgE-mediated triggering of RBL-2H3 cells. Materialsand Methods:Secretoryresponse of the cells was monitored via the released B-hexoseaminidas activity; tyrosine phosphoryiation of cellular proteins was followed by immunoprecipitation; changes in the concentration of intracellular free Ca2+ was monitored using Flue-MM. To identify the C3a-binding ceil-membrane molecule radiolabelled C3a was cross-linked to the cell surface by the membrane-impermeable homobifuncional cmsslinker, BS3. Reeultr: C3a inhibited IgE-mediated triggering of RBL-2H3 cells dose dependentfy while C6a had no effect at all. When added alone, none of the complement-peptides biggered the secretory response of these muoosal-type cells. All the s&ps of the signaling pathway studied were inhibited dosedependently by C3a: protein phosphoiylation, hydrolysis of phosphatidyl inositides, transient rise in free cytosolic Ca2+-ion concentration. The results of ligand crosslinking experiments revealed a C3a-binding molecule of appr. 30 kD. As a result of immunoprecipitation of the lysates with anti-C3a followed by Westem-blotting with an antibody specific for the B-chain of FcsRI (kindly provided by Dr.J.-P. Kinet) a molecule of similar size could be identified. Conclusion: Our results reveal a novel function of C&I: its inhibitory capacity of the IgE mediated triggering of mucosal mast cells, most probably mediated via its interaction with the B-chain of FcERI. This inhibition takes place at a C3a concentration-range orders of magnitude lower than those where this anaphylatoxic peptide exerts its secretory stimulus to serosal type cells. Since inhaled allergens may activate the complement system at mumsal surfaces and consequently C3a peptides may be generated locally, our finding may also have a relevance to the in vivo occuring situation.

P.5.13.02

Haemocarboperfusion as a method of bronchial asthma immunotherapy ._

G.I. Lisenko, O.B. Yashchenko, S.V. Daniluk. Kiev Medical Academy of post-graduate educafion, Ukraine Study on the bronchial asthma at an angle of autoimmuny pathology helped reveal an important mle of low molecular immune suppressive lymphopeptide with mol. wgt. 300-500 daltons and autoantibodies against DNA-ase enzyme in the mechanism of formation of secondary immune suppression. The use of blood perfusion through haemosorbents in the patient with bronchial asthma in order to eliminate disturbance of the humoral regulation of immunocompetent cells and the restraint of action of the above factors resulted in the development of compensatory-humoral reactions, which normal&d indices of immune homeostasis. There was an increase in the content and functional activity of T-lymphocytes, the number of “active” T-lymphocytes, and the number and-activily of T-suppressory cells capable to limit the autoantibody-formation The immune stimulating effect of haemocarboperfusion may be utilised during treatment of patients with bronchial asthma in cases of development of immune deficient condition, low efficacious action of glucocottlwids and cytostatics, and development of corticusteroids dependency and wtticostemids resistance.

P.5.13.03

Early immunologic ef?ectsof rush Immunotherapy with Dennatophagoides ptwvnyssinus In asthmatlc persons

S. Paranos ‘, S. Petrovic *. ’ Department of Allergy and Clinical Immunol, Uniwsiiy Medical Center Zvezdara, Belgrade, Yugoslavia,2 Vimfogy and lmmunobiology Institute Torlak, Belgrade, Yugoslavia

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P.5.13

Immunotherapy of allergy

P.5.13.01

IgE-induced triggering of RBL-2H3 cells is inhibited by complement-peptide C3a but not by C5a

A. Erdei, I. Pecht ‘. Department of Immunology, E&v& L. University, Gad, Hungary, ’ The Weizmann Institute of Science, Rehovot, Israel Introduction: Mast cells and basophils are known to be triggered by allergens that clusters their high affinity IgE-binding receptors, FcsRI. The anaphylatoxic activity of complementderived peptides C3a and C5a has been known for a long time; however, semsal and mucosal type mast cells are known to respond differently to these stimuli. Namely, only serosal type cells are activated by peptidergic stimuli, mucosal type cells, including the rat cell line RBL-2H3 are

Introduotlon: Allergen specific immunotherapy (IT) is one of therapeutic modality for susceptible younger persons with mild asthma. Materials and Methods: We estimated effects of Dot rush-immunotheram in comparison with placebo treatment in fourteen highly Dpt-susceptible ad& (9 females, 5 males, mean age 29.6). They were randomized in two groups (immunotherapy and placebo) and treated in a single blind manner. To minimize side-effects we modified the protocol by adjusting allergen doses to each patient separately. That way patients could receive low, middle or high cummulative dose of allergen over 4 month of immunotherapy, depending on the initial allergen concentration. Immunologic (total and Dpt-specific serum IgE and IgG antibodies/EIA, Pharmacia/) and clinical parameters (spirometry, medication score and skin testing) were registered before treatment, after 2 weeks, at the 2nd month and after 4 months of Immunotherapy onset. Results: At the end of 4 months two patients reached high cummulative dose, four gained middle and one person reached low cummulative dose of allergen. None of them had life-threatening side effects overtreatment. Obtained results revealed significant influence of immunotherapy on Dpt-specific serum IgG synthesis (Kruskal Wallis, p < 0.05) and on the late phase skin reaction with Dpt (U test, p < 0.05). The omission of significance for total and Dpt-specific