Bone marrow samples from 55 patients with multiple myeloma (MM) and 23 patients with monoclonal gammopa- thy of undetermined significance (MGUS) were ...
Plasma Cells in Multiple Myeloma Express a Natural Killer Cell-Associated Antigen: CD56 (NKH-1; Leu-19) By Benjamin Van Camp, Brian G.M. Durie, Catherine Spier, Marc De Waele, Ivan Van Riet, Elizabeth Vela, Yvette Frutiger, Lynne Richter, and Thomas M. Grogan Bone marrow samples from 55 patients with multiple myeloma (MM)and 23 patients with monoclonal gammopathy of undetermined significance (MGUS) were evaluated with a broad panel of monoclonal antibodies. Plasma cells from 78% (43/55) of patients with MM strongly expressed the natural killer cell antigen CD56 (NKH-1, Leu-19). Of the 23 patients with MGUS, none showed strong CD56 reactivity, although three had weak reactivity in less than 20% of plasma cells. Myeloma cells expressing CD56 did not coexpress the CD57 or CD16 antigens. Patients with CD56-positive plasma cells had both indolent and aggres-
sive disease. However, the 12 CD56-negative patients had predominantly aggressive disease with an unexpected preponderance of K Bence Jones only myeloma (5/10 [50%] evaluable patients). Polyclonal plasma cells from nonneoplastictissue sites (normal bone marrows, lymph nodes, tonsillar biopsies, and gut-mucosa biopsies)showed a near absence of CD56. We conclude that isolated, strong CD56 expression is common in MM, but not in MGUS or reactive plasma cells. The potential biologic importance of CD56 positivity in myeloma is reviewed. 0 1990 b y The American S o c i e t y of Hematology.
T
acetone at 4OC for 10 minutes and air-dried at room temperature. Mouse monoclonal antibody (MoAb) was applied for 30 minutes to detect human cell surface (and/or cytoplasmic) antigen. Negative controls consisted either of mouse ascitic fluid (Bethesda Research Laboratories, Gaithersburg, MD) or irrelevant isotypic-matched antibody instead of the primary antibody. The first stage included MoAbs to the following antigens (cluster designations [CD] follow in brackets): NK antigens Leu-7 [CD57], Leu-1 l b [CD16], and Leu-19 [CD56] (Becton-Dickinson, Mountain View, CA); B antigens: B1 [CD20], B2 [CD21], B4 [CDlO] (Coulter Immunology, Hialeah, FL); Leu-14 [CD22] (BectonDickinson); immunoglobulins ( K , X, p, 7 , 6, a;Becton-Dickinson); plasma cell antigens: PCA-1 (Coulter); PC-1 (supplied by Drs Kenneth Anderson and Lee Nadler, Dana-Farber Cancer Institute, Boston, MA); Leu-17 [CD38] Becton-Dickinson); CALLA [CDlO], HLA-DR and T-cell antigens: Leu-1 through Leu-9 [CD1-5,7,8]; human progenitor cell antigen (HPCA) [CD34], transferrin receptor [CD71], myelomonocytic antigens (LMl [CDlS], LM3, LM5) (all obtained from Becton-Dickinson);myeloid antigen MY7 [CD13], and anti-interleukin-2 receptor (anti-IL-2R) [CD25] (Coulter). Anti-leukocyte common antigen [CD45] was obtained from Dakopatts, Copenhagen, Denmark. Plasma cells were scored as strongly or weakly positive in comparison with negative controls. More than 100 plasma cells per
HE CURRENT REPORT resulted from an unexpected observation during the analysis of immunophenotyping of bone marrow samples from patients with multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS). The planned study was an analysis of the phenotypic pattern of lymphocytes, natural killer cells, and monocyte/macrophages in myeloma bone marrows as a basis for the potential use of interleukin-2 (IL-2) and related forms of immunotherapy’-’ in patients with plasma cell dyscrasias. The test panel included antibodies against natural killer (NK) cells, since they represent an important component of lymphokine activated cell^.^-^ N K cell antibodies tested included anti-CD57, -CD11b, -CD16, and -CD56.’-’’ Unexpectedly, plasma cells from patients with MM were found to be strongly positive for CD56. A detailed analysis was then conducted comparing MM, MGUS, and normal, polyclonal plasma cells from different tissue sites. This report documents the results of this analysis and evaluates the potential biologic significance of this phenotypic aberrancy in multiple myeloma. MATERIALS AND METHODS
Patients
Consecutively accrued bone marrow aspirates of 78 patients with monoclonal gammopathy were processed for immunotyping. The final diagnosis at the time of bone marrow sampling was defined according to the Southwest Oncology Group (SWOG) criteria.” Fifty-five patients had MM, and 23 had MGUS. Twenty-four MM patients were studied before any treatment, while the others had progressive (21 patients) or refractory (10 patients) disease. All isotypes of monoclonal immunoglobulins (Ig) were detected among these patients. The bone marrows of 14 patients with nonhematologic diseases, as well as eight with reactive lymph node plasmacytosis, three with tonsillar plasmacytosis, and four with inflammatory gastrointestinal tract biopsies (with a component of plasmacytosis), were examined for CD56 positivity of plasma cells. Immunologic Marker Studies Immunoperoxidase typing. Snap-frozen marrow clot section and core biopsies, as well as cytccentrifuge (Shandon Southern Instruments, Sewickley, PA) slide preparations of the myeloma cell material for immunologic evaluation, were used as previously reported.” Biotin-avidin conjugates and horseradish peroxidase labeled with diaminobenzidine tetrahydrochloride (DAB) as the detection agent were used. Cytocentrifuge slides were fixed in Blood, Vol 76,No 2 (July 15). 1990:pp 377-382
From the Section of Haematology/Oncology. Department of Internal Medicine and the Arizona Cancer Center, University of Arizona College of Medicine. Tucson, AZ; the Department of Pathology, University of Arizona Medical Center. Tucson, AZ; the Division of HaematologyIImmunology, Faculty of Medicine, Free University of Brussels ( W B ) , Brussels, Belgium: and the Department of Haematology. University of London. Charing Cross Hospital, London, UK. Submitted August 29, 1989: accepted March 19, 1990. Supported in part by Grant No. 3.0033.88from the National Foundation of Scientific Research (NFWO), Belgium, by Grant CA17094 from the National Cancer Institute. and by donations to the Myeloma Research Fund at the University of Arizona. Address reprints requests to Brian G.M. Durie. MD, Department of Haematology. University of London, Charing Cross Hospital. Fulham Palace Rd, London. UK W6 8RF. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C.section 1734 solely to indicate this fact. 0 1990 by The American Society of Hematology. 0006-4971/90/7602-0OI 5$3.00/0 377
378
VAN CAMP ET AL
preparation were screened and evaluated in terms of both number and degree of positivity. Immunofluorescence typing: Flow cytometry. With a previously described method, antigens were detected by flow cytometric analysis using indirect immunofluorescence (IF).’* Briefly, cultured myeloma cells were washed in RPMI medium; trypan blue dye exclusion was used to establish viability. The myeloma cells were then incubated with the same battery of MoAbs as before for 30 minutes at 4°C. A second stage goat antimouse IgG fluorescein isothiocyanate (FITC) conjugate was applied and incubated as above. After cold phosphate-buffered saline (PBS) washing, the cells were kept on ice in the dark and analyzed immediately. Flow cytometry was conducted with fluorescent antibodies analyzed by a Coulter EPICS V flow cytometer (EPICS Division, Coulter Immunology, Hialeah, FL) as previously described.” Data obtained by flow cytometry were analyzed by using MDADS (Multiparameter Data Acquisition and Display System) “Immuno” program; the coplot display of composite histograms was created by using the EASY 88 (Extended Analysis System with Intel 80/99 Microprocessor) analysissystem. Two-color immunofluorescence microscopy. Simultaneous detection of cytoplasmic CD56 and immunoglobulin (Ig) in plasma cells was carried out using cell suspensions of bone marrow (fixed in acetone) and following a previously described method.13An antigenpositive plamsa cell was scored as strongly or weakly positive, and the percentage of positive plasma cells was calculated. A minimum of 100 plasma cells were assessed. Patients classified as CD56positive had more than 50% positive plasma cells. Simultaneous staining of surface CD56 and intracytoplasmic Ig was carried out using viable cells (without fixative) that were first incubated with anti-CD56 followed by anti-mouse Ig-TRITC. The suspension was then cytocentrifuged on slides, fixed, stained with antihuman-Ig-FITC, and evaluated using two color fluorescent microscopy as previously de~cribed.’~ Immunogold-silver staining procedure. The immunogold-silver staining procedure was done as described e1~ewhere.l~ Cytocentrifuge preparations were fixed in phosphate-buffered 9.25% formaldehyde, 45% acetone, pH 6.6 room temperature. They were rinsed twice with distilled water, and then silver enhancement was done with the Intena TJ silver enhancement kit (Janssen Biotech, Olm, Belgium) for 30 minutes at 36OC. This was prepared following the recommendations of the manufacturer.Afterwards, the preparations were rinsed twice with distilled water and counterstained with May-Grunwald-Giemsa.They were mounted in DPX mount material (BDH, Montreal, Canada) and examined by bright field light microscopy. Plasma cells were called positive if more than five granules were visible at the surface membrane. Patients with more than 50% of CD56-positive plasma cells were accepted as having CD56-positive MM (see cover photograph). 51 Chromium release cytotoxicity assay. CD56-positive myeloma cells (8226 cell line and a direct patient sample) were tested in the chromium release assay using standard methodology.’ The NK sensitive myeloid leukemia cell line K562 and the NK resistant lymphoma cell line Daudi were used as targets. Complete details are part of a separate publication.’6 RESULTS
Expression of CD56 on Plasma Cells
Plasma cells from patients with MM, MGUS, and normal tissues were evaluated. The overall CD56 positivity of plasma cells is summarized in Table 1. Of 55 patients with multiple myeloma, 43 (78%) were positive. Using the same
Table 1. CD56-Positive Plasma Cells
MM MGUS Normal tissues Bone marrows Lymph nodes GI biopsies Tonsil biopsies
No.
%
43/57
?a
0123
0
Oil0
0 0 0 0
oia 014
013
Positivity defined as greater than 50% plasma cells positive with strong antigen reactivity. See discussion in text.
criteria (greater than 50% cells reactive), there were no MGUS-positive patients. Likewise, no normal tissues showed strong reactivity. A few specimens showed weak reactivity, particularly from patients with MGUS. Two patients with MGUS had detectable although weak CD56 plasma cell reactivity using immunoperoxidase. Three had less than 20% plasma cells with weak CD56 positivity using surface immunofluorescence. Samples from patients with active M M were the only ones showing a strong pattern of reactivity. Studies were conducted to assess surface plus cytoplasmic expression of CD56 using acetone fixation to allow membrane permeabilization for detection of intracellular CD56 (Fig 1). The four methods gave comparable results: immunoperoxidase: cytoplasmic + surface expression, 8 1% CD56-positive (22/27); immunofluorescence: cytoplasmic expression, 80% (8/ 10); immunofluorescence: surface expression, 72% (1 3/ 18); immunogold (see cover photograph): cytoplasmic + surface expression, 72% (1 3/ 18). Twenty-nine patients with non-hematologic diseases were investigated for the expression of CD56 in plasma cells, using the various listed methods. In 14 bone marrow samples, only weak positivity was observed in a minority (less than 10%) of the plasma cells. No strong positivity was detected. In addition, the polyclonal plasma cells of eight reactive lymph nodes, four gastrointestinal biopsies, and three tonsillar biopsies studied by the peroxidase techniques were completely negative for CD56. Documentation of Co-Expression of CD56 and CD38 Using Flow Cytometry
The specific expression of CD56 on plasma cells was documented by assessment of the dual co-expression of CD38 and CD56 using flow cytometry. Results are shown in Fig 2. Panel A shows results with a direct (patient) bone marrow sample replaced with plasma cells. There was greater than 90% co-expression of CD56 and CD38. In panel B, results with the 8226 myeloma cell line are shown. There was 51% co-expression of CD56 and CD38; expression of CD56 in approximately half of the cells in this well-studied myeloma cell line has been previously reported and is discussed later in the text. Further assessment of a drug resistant 8226 myeloma line (8226 Dox 40) showed a comparable heterogeneous expression with both CD38positive and -negative cells in the context of full CD56 expression.
NK ANTIGENS (CD56) IN MULTIPLE MYELOMA
379
Fig 1. Demonstration of CD56 (L19) positivity by the immunoperoxidasetechnique. The eight panels in this photomicrograph show the morphologic and phenotypic features of a patient whose plasma cells showed clear CD56 positivity plus co-expression of K (k), Leu-17 (L17). and Ki-67 (K67). The obvious plasma cell morphology is combined with the monoclonalK positivity and CD38 positivity, confirming a plasma cell phenotype. Note the absent X, but strong Ki67 (K67). positivity. (Cytospin immunoperoxidase preparation with original magnification of ~ 6 3 0 . 1
Evaluation of CD56 Expression in Myeloma and Clinical Disease Forty-three (78%) patients with MM had more than 50% CD56-positive plasma cells. The CD56-positive patients had different stages and disease activity: 17 were newly diagnosed, 18 had progressive disease, and eight were completely refractory to treatment (Fig 3). At the time of analysis, all MM patients had stage IIIA disease, except for three with stage IIIB and three with stage IA disease. In addition, patients with poor prognostic features (such as high /3-2 microglobulin, high labeling index, p-glycoprotein positivity, and plasmablastic morphology), as well as good prognostic features (eg, smoldering myeloma or indolent myeloma), were all found to have CD56-positive plasma cells. All types of monoclonal Igs were present (20 IgG, 9 IgA, 4 K (only), 1 X (only), 1 I@). Additional Antigen Expression in CD56-Positive Myeloma Fig 2. Flow cytometry evaluation of a patient bone marrow sample (A) and the 8226 X 8ence Jones myeloma cell line (8).The patient specimen shows remarkable (greater than 90%) coexpression of CD38 (Leu-17) and CD56 (Leu-19). The 8226 culture shows 51% CD56 (Leu-19) expression in this Leu-17 (CD38)positive plasma cell line.
Expression of other leukocyte markers on plasma cells was also detected in 13 of 22 cases (59%) evaluated (Table 2). In 6 of 22 CD56-positive MM cases tested, there was additional expression of B cell markers CD19 and/or CD22. Two patients had C D 10 (CALLA)-positive plasma cells, and three patients were found to have class I1 histocompatibility
380
%
VAN CAMP ET AL
100
n*nP
80 60
0
* %
40
0 0 0
no t h e r a p y MM p r o g r e s s i v e MM r e f r a c t o r y MM MGUS NL
0
20 0
MGUS
MM
NL
antigen expression. In two patients, myelomonocytic antigens were found (CD15, LM3). The plasma cell antigen-I (PCA-I) monoclonal antibody was negative in 15 of 24 cases and only weakly expressed in 9 of 24 cases, whereas CD38 was strongly positive on plasma cells from all the patients with M M and MGUS. Of 12 M M patients with CD56negative plasma cells, 4 were previously untreated, and the others had progressive or refractory disease. Of considerable interest, 5 of IO patients had K light chain only MM. Of five tested, four had additional expression of B cell markers.
N K Functional Activity of CDSbPositive Myeloma Cells Bone marrow myeloma cells from five patients with strongly CD56-positive myeloma cells were tested in a chromium release assay for cytotoxicity. N o significant cytotoxicity was observed (-2% to + 5.4%) with the K562 or Daudi cell lines as target cells. The 8226 (CD56-positive) myeloma cell line was also tested, both with and without stimulation using IL-2 plus lymphokine-activated supernatant. No evidence of cytotoxicity was observed. Of note, great care was taken to be sure that CD56 positivity was fully retained at the time of testing of both the patient and cell line samples. It was observed that surface expression could be lost with even brief in vitro liquid culture. Patients with loss of positivity were excluded from these specific functional, as well as flow cytometric, analyses. Table 2. Additional Antigen Expression on Plasma Cells in Patients With CD56-Positive Plasma Cells
Overall additional antigen expression Specific antigens B antigens (CD32, CD 19) la (HLA-DR) CALLA (CD 10) Myelomonocytic (CDl5, Leu-M3) T antigens (CD8) IL-2R (CD25)
13/22
59
6/22 3/22 2/22 2/22 2/22 1/22
27 14 9
9 9 5
Some plasma cells expressed greater than one additional antigen: eg, myelomonocytic cases also expressed la, and a CD8-positive case expressed CD22.
Fig 3. The percent of CD56positive plasma cells in MM, MGUS, and in normal or nonhematologic conditions (NL). The MM patients are separated into those having received no therapy, those with progressive disease, and those refractory to treatment.
DISCUSSION
In this study we have documented the frequent occurrence of the N K cell-associated marker CD56 on plasma cells in bone marrow specimens of patients with MM, but not with MGUS or non-hematologic diseases. Reactive plasma cells in lymph nodes, tonsil, and gut were uniformly CD56negative. In addition, other N K cell markers, such as CD57 and CD16, were notably absent from myeloma cells. However, additional expression of B cell, CALLA, myelomonocytic, or other leukocyte antigen^''.^^ occurred in 59% of the CD56-positive myeloma patients. The CD56 antigen recognized by the monoclonal antibodies NKH-1 and Leu-19 is a 174 to 185 Kd glycoprotein that is expressed on essentially all human NK cells and the subset of T lymphocytes and IL-2-activated thymocytes that mediate major histocompatibility (MHC) unrestricted cytotoxi ~ i t y . ~However, ' the expression of CD56 is not restricted to these cell types; it has been described on certain CD4-positive T helper cells, myeloid leukemia cells, the KGla immature hemopoietic cell line, trophoblast cells, small cell lung carcinoma, and even urchin coelomonocytes, and, finally, on neural t i s s ~ e s . ~ The ' . ~ *fact that this antigen appears on such disparate cell types suggests a basic and evolutionarily conserved role. In this respect, the recent finding that the CD56 antigen is identical to the neural cell adhesion molecule N-CAM is of considerable interest.33Adhesion molecules such as N-CAM have been implicated in cell migration, embryonic development, and cellular adhesion. Therefore, it is conceivable that the expression of CD56 on myeloma cells may be relevant to the biology of the disease. Considering the potential significance of CD56 expression on myeloma cells, several additional points are noteworthy. First, the other N K cell markers CD57 and CD16 were absent. There was also no indication that the CD56-positive myeloma cell had N K functional activity. This implies that the isolated CD56 expression is aberrant in some way, rather than of true functional significance. On the other hand, since the CD56 molecule has homology with the neural cell adhesion molecule N-CAM, the expression of CD56 may
38 1
NK ANTIGENS (CD56) IN MULTIPLE MYELOMA
contribute t o the specific biologic behavior of t h e myeloma in individual patients (eg, presence or absence of plasma cell leukemia). In this regard, it is of interest that although t h e CD56-positive group of patients contains patients with both indolent a n d aggressive disease, the absence of C D 5 6 expression occurred in patients with an unexpected preponderance of K Bence Jones only myeloma and predominantly aggressive disease (one with plasma cell leukemia). This may relate to recent observations of the expression or non-expression of adhesion molecules in other hematopoietic malignant disease, such as non-Hodgkin’s lymphomata and chronic myelogenous leukemia, in association with particular patterns of disease activity.34 Whatever proves to be the underlying biologic significance of CD56 expression, this report documenting the frequent expression of CD56 in myeloma is important for a variety of reasons. First, strong C D 5 6 expression is a reliable discriminator between myeloma and MGUS. No strong CD56 expression was noted in patients with MGUS. Second, the
CD56 marker is a useful target for purging myeloma cells from bone marrow specimens, and a recent preliminary report documented successful purging.35 Third, the CD56 positivity in myeloma can be used as a basis for further exploring t h e role of adhesion molecules, particularly the interaction between malignant plasma cells and other bone marrow cells including stromal cells, which have recently been noted to express N - C A M molecules.36 The apparent tropism of myeloma cells for areas of active bone marrow activity may relate t o expression of adhesion molecules on bone marrow accessory cells or other hematopoietic elements. Clearly, this initial observation of CD56 positivity in myeloma is believed to warrant more detailed evaluation and m a y lead to further findings of both clinical and biologic significance. NOTE
Please note that CD56 positivity as shown by the immunogold technique is the cover illustration for this issue of Blood.
REFERENCES 1. Rosenberg SA, Lotze MT, Muul LM, Chang AE, Avis FP,
Leitman S, Linehan WM, Robertson CN, Lee RE, Rubin JT, Seipp CA, Simpson CG, White DE: A progress report on the treatment of 157 patients with advanced cancer using lymphokine-activated killer cells and interleukin-2 or high dose interleukin-2 alone. N Engl J Med 316:889,1987 2. Adler A, Chervenick PA, Whiteside TL, Lotzova E, Herberman RB: Interleukin 2 induction of lymphokine-activated killer (LAK) activity in the peripheral blood and bone marrow of acute leukemia patients. I. Feasibility of LAK generation in adult patients with active disease and in remission. Blood 71:709, 1988 3. Topalian SL, Solomon S, Avis FP, Chang AE, Freerksen DL, Linehan WM, Lotze MT, Robertson CN, Seipp CA, Simon P, Simpson CG, Rosenberg SA: Immunotherapy of patients with advanced cancer using tumor infiltrating lymphocytes and recombinant interleukin-2: A pilot study. J Clin Oncol6:839, 1988 4. Heo DS, Whiteside TL, Kanbour A, Herberman RB: Lymphocytes infiltrating human ovarian tumors. I. Role of leu-19 (NKH1)positive recombinant IL2-activated cultures of lymphocytes infiltrating human ovarian tumors. J Immunol 140:4042, 1988 5. Topalian SL, Muul LM, Solomon D, Rosenberg SA: Expansion of human tumor infiltrating lymphocytes for use in immunotherapy trials. J Immunol Methods 102:127, 1987 6. Whiteside TL, Heo DS, Takagi S, Herberman RB: Characterization of novel anti-tumour effector cells in long term cultures of human tumour infiltrating lymphocytes. Transplant Proc 20:347, 1988 7. Abo T, Balch DM: Differentiation antigen of human NK and K cells identified by a monoclonal antibody (HNKl). J Immunol 127:1024, 1981 8. Lanier LL, Le AM, Phillips JH, Warner NL, Babcock GF: Subpopulations of human natural killer cells defined by expression of the leu-7 (HNK-1) and leu-I1 (NK-15) antigens. J Immunol 131:1789,1983 9. Hercend T, Griffin JD, Bensussan A, Schmidt RE, Edson MA, Brennan A, Murray C, Daley JF, Schlossman SF, Ritz J: Generation of monoclonal antibodies to a human natural killer clone: Characterization of two natural killer associated antigens, NKH 1A and NKH2, expressed on subsets of large granular lymphocytes. J Clin Invest 75:932, 1985 10. Lanier LL, Le AM, Civin CI, Loken MR, Phillips JH: The
relationship of CD16 (Leu-11) and Leu-I9 (NKH-1) antigen expression on human peripheral blood NK cells and cytotoxic T lymphocytes. J Immunol 136:4480, 1986 11. Durie BGM: Staging and kinetics of multiple myeloma. Semin Oncol13:300,1986 12. Grogan TH, Durie BGM, Lomen C, Splier C, Wiat DP, Nagle R, Wilson GS, Richter L, Vela E, Maxey V, McDaniel K, Range1 C: Delineation of a novel pre-B cell component in plasma cell myeloma: Immunochemical, immunophenotypic, genotypic cytologic, cell culture, and kinetic features. Blood 70:932, 1987 13. Van Camp B, Reynaert P, Broodtaerts L: Studies on the origin or the precursor cells in multiple myeloma, macroglobulinemia of Waldenstrom and benign monoclonal gammopathy. I. Cytoplasmic isotype and idiotype distribution. Clin Exp Immunol 44:82, 1981 14. Van Camp B, Thielemans K, Dehou MF, De Mey J, De Waele M: Two monoclonal antibodies (OKTa and OKT10) for study of the final B cell maturation. J Clin Immunol2:67, 1982 15. De Waele M, De Mey J, Renmans W, Labeur C, Jochmans K, Van Camp B: Potential of immunogold-silver staining for the study of leukocyte subpopulations as defined by monoclonal antibodies. J Histochem Cytochem 34:1257, 1986 16. Mills D, Durie BGM, Grogan TM, Vela E, Frutiger Y: Expansion of two subpopulations ( C D 8 + , C D 1 1 + and CD3 - CD56+) of tumour infiltrating lymphocytes with cytotoxic activity in multiple myeloma bone marrow. (submitted) 17. Anderson KC, Bates MP, Slaughenhoupt B, Schlossman SF, Nadler LM: A monoclonal antibody with reactivity restricted to normal and neoplastic cells. J Immunol 132:3172, 1984 18. Anderson KC, Park EK, Bates MP, Leonard RCF, Hardy R, Schlossman SF, Nadler LM: Antigens on human plasma cells identified by monoclonal antibodies. J Immunol 130:1132, 1983 19. Ling NR, MacLennan ICM, Mason DY: B cell and plasma cell antigens: New and previously defined clusters, in McMichael AJ, Berverley PCL, Cobbold S, Crumpton M, Gilks W, Gotch FM, Hogg N, Norton M, Ling N, MacLennan ICM, Mason DY, Milotoin C, Epiogo L, Halter D, Waldman H (eds): Leukocyte Typing 111: White Cell Differentiation Antigens, Chapter 7. Oxford, UK, Oxford University Press, 1987 20. San Miguel JF, Caballero MD, Gonzales M, Zola H, LopezBorrasco A: Immunologic phenotype of neoplasms involving the B cell in the last step of differentiation. Br J Haematol62:75, 1986
382
21. Caligaris-Cappio F, Bergui L, Tesio L, Pizzolo G, Malavasi F, Chilosi M, Campana D, Van Camp B, Janossy G: Identification of malignant plasma cell precursors in the bone marrow of multiple myeloma. J Clin Invest 76:1242, 1985 22. Durie BGM, Grogan TM: CALLA-positive myeloma: An aggressive subtype with poor survival. Blood 666:229, 1985 23. Spier C, Durie BGM, Grogan TM, Richter LC, Vela E, Frutiger Y, Ranger CS: T cell antigen positive multiple myeloma. Mod Pathol (in press) 24. Durie BGM, Grogan TM, Spier C, Vela E, Baum V, Rodriquez MA, Fruitger Y: Myelomonocytic myeloma cell line (LB84-1). Blood 73~770,1989 25. Kubagawa H, Vogler L, Capra JHD, Conrad MF, Lawton A, Cooper MD: Studies on the clonal origin of multiple myeloma. J Exp Med 150:792,1979 26. Brown G, Bruce CM, Howie AJ, Lord JM: Stochastic or ordered lineage commitment during haemopoiesis. Leukemia 1:150, 1987 (editorial) 27. Duperray C, Klein B, Durie BGM, Zhang X, Jourdan M, Poncelet P, Favier F, Vincent C, Brochier J, Lenoir G, Bataille R: Phenotypic analysis of human myeloma cell lines. Blood 73556, 1989 28. Timonen T, Ortaldo J, Herberman RB: Characteristics of human large granular lymphocytes and relationship to natural killer and Kcells. J Exp Med 153:159, 1981 29. Uchida A, Yagita M, Sugiyama H, Hoshino T, Moore M: Strong natural killer (NK) cell activity in bone marrow of myeloma patients: Accelerated maturation of bone marrow NK cells and their interaction with other bone marrow cells. Int J Cancer 34:375, 1984 30. Lanier LL, Le AM, Ding A, Evans EL, Krensky AM, Clayberger C, Phillips JH: Expression of Leu-19 (NKH-1) antigen on IL2-dependent cytotoxic and non-cytotoxic T cell lines. J Immuno1 138:2019, 1987
VAN CAMP ET AL
31. Barkowitz RS, Umpierre SA, Goldstein DP, Anderson DJ: Cross-reactivity of monoclonal antibodies, directed against lymphocyte markers with trophoblast cells of normal placenta and gestational placenta, hydatidiform mole and gestational choriocarcinoma. Gynaecol Oncol29:94, 1988 32. Koros AMC, Taylor MJ, Hiserodt HC, Stephens B, Lakomy RJ, Dalbow M, Raikow R, Lange J, Wagner M: Sea urchin coelomonocytes, human peripheral blood NK cells, and human small-cell lung cancer cells share evolutionarily conserved antigens HNK-I (Leu-7) and N901 (NKH-I), in McMichael AJ, Beverly PCL, Cobbold, Crumpton MJ, Gilks W, Gotch FM, Hogg N, Horton M, Ling N, MacLennan ICM, Mason DY, Milstein C, Spiegelhalt D, Waldman H (eds): Leukocyte Typing 111: White Cell Differentiation Antigens, Chapter 16. Oxford, UK, Oxford University Press, 1987, p 866 33. Lanier LL, Testis R, Bindl J, Phillips JH: Identity of Leu-19 (CD56) leucocyte differentiation antigen and neural cell adhesion molecule. J Exp Med 169:2233, 1989 34. Stauder R, Greil R, Schulz TF, Thaler J, Gattringer C, Radaskiewicz T, Dierich MP, Huber H: Expression of leucocyte function-associated antigen-1 and 7F7-antigen, an adhesion molecule related to intercellular adhesion molecule-1 (ICAM-1) in non-Hodgkin’s lymphomas and leukaemias: Possible influence on growth pattern and leukaemic behaviour. Clin Exp Immunol77:234, 1989 35. Van Riet I, Durie B, Grogan T, Van Camp B: A human natural killer cell antigen (Leu-19) NKH-I is expressed by myeloma cells and can be used for bone marrow purging. Bone Marrow Transplant 4:105, 1989 (suppl2) 36. Thomas PS, Pietrangeli CE, Hayashi S, Schachner M, Goridis C, Low M, Kincade PW: Stromal cells that support lymphopoiesis. Leukemia 3: 171, 1988
