Leukemia Re,search Vol 6. N o 1, pp 123 125, 1982 t)145-2126/82/010123-03503.00/0 Primed in Grea! Britain Pergamon Press Ltd.

BRIEF COMMUNICATION

B I N D I N G O F P E A N U T A G G L U T I N I N T O

N O R M A L H U M A N L Y M P H O C Y T E S A N D

T O L E U K E M I C C E L L S *

MOSHE BARZILAY,t ESTHER ROSENTHAL,~ GEORGE KENDE,+ + YAIR REISNER,~" NATHAN SHARON't" and BRACHA RAMOT +

+Department of Biophysics. The Weizmann Institute of Science, Rehovot, Israel; ++Department of Hematology, Chaim Sheba Medical Center and

Tel Aviv University Sackler Medical School, Israel

(Received 24 September 1981. Accepted 12 October 1981)

Key words: Peanut agglutinin, human lymphocytes, acute leukemia, chronic leukemia."

PREVIOUS WORK in our laboratories has demonstrated that peanut agglutinin (PNA), a lectin specific for the disaccharide o-galactose-fll---. 3-N-acetyl-D-galactosamine, binds exclusively to immunologically immature lymphocytes in murine thymus [7], fetal liver [-6], bone marrow and spleen [8], and that on the mature cells the PNA receptor is masked by sialic acid. It was further found that human thymocytes can be fractionated by selective agglutination with PNA into two subpopulations: an agglutinated, PNA ÷ fraction, which exhibits considerably lower immune-reactivity than the unagglutinated, PNA- fraction, which binds the lectin poorly [9]. Other laboratories have subsequently reported similar observations both for murine lymphocytes and for human thymocytes [1,4; reviewed in 12 for more recent reports see 10, 11]. The fact that over 9030 of the immature PNA + human thymocytes bear the receptor for sheep red blood cells, has led to the suggestion that the masking of the receptor for this lectin by sialic acid occurs at a later stage of T cell maturation than the expression of sheep red blood cell receptor 1-93. The lectin bound to less than 2°0 of the peripheral blood lymphocytes from normal donors [9]. Upon treatment of the cells with sialidase, binding occurred to all cells; both B and T lymphocytes were reported to have on the average 3.8 x 10 6 PNA binding sites per cell [5].

Since in normal human peripheral blood lymphocytes the percentage of PNA + cells is very low. an increase in the number of such cells could be an indication of a disturbance in cell maturation. A survey of a small number of leukemic patients has indeed shown increased binding of PNA to cells from certain types of leukemia [9]. This communi- cation presents the results of a survey of PNA binding to peripheral blood mononuclear cells from about 220 leukemic patients seen during the last five years at the Chaim Sheba Medical Center. Ramat Gan. Israel.

Mononuclear cells were isolated from the peripheral blood of leukemic patients as well as from 24 normal donors, by centrifugation over Ficoll-Hypaque [2]. Binding of PNA

* This study' ,aas supported by National Institutes of Health Contract NOI-CB-74163 {to N.S.} and by' the Gershon Meerboim Foundation (to B.R.I.

Correspomtencc to: Moshe Barzilay. Department of Biophysics. The Weizmann Institute of Science, Rehovot 76100. Israel.

123

124 MOSHE BARZ1LAY et al.

TABLE 1. PNA* CELLS IN PERIPHERAL BLOOD LYMPHOCYTES OF PATIENTS WITH DIFFERENT TYPES OF LEUKEMIA AND OF NORMAL INDIVIDUALS

Disease

Number Number of patients with of different percentages of

patients PNA + cells examined < 2 2-19 20-49 50-79 80--100

Acute leukemias, total 128 47 19 9 12 41 Lymphocytic 85 36 11 4 8 26 Myelo-monocytic 29 I0 5 2 3 9 Burkitt 14 1 3 3 1 6

CLL 68 40 16 3 3 6 ALL in remission 28 11 14 l I 1 Normal 24 24 0 0 0 0

No correlation could be found between the number of blast cells in the peripheral blood of individual leukemic patients and the binding of PNA to cells. The number of blasts in acute leukemia varied between 30 and 100°~,, with the majority of cases showing more than 75°o blasts. In DLL, ALL in remission and in normal peripheral blood, no blasts were found.

to the cells was monitored as previously described I-9], using fluorescein isothiocyanate- PNA (final concentration 250 ~g/ml; 3-5 × 106 cells in 0.2 ml final volume).

Pathologic cells of patients with different types of leukemia varied in their PNA binding properties (Table 1). A considerable number of patients had a high level ( > 20°,0) of PNA ÷ cells. We classify such patients as lectin-positive, whereas those with lower levels of lectin binding cells are classified as lectin-negative. Forty-five per cent of patients with ALL and 48~o of patients with myeloid leukemia, were lectin-positive; 71°o of Burkitt's leukemia and lymphoma were also found to have high proportions of PNA + cells. Of special significance is the finding that 11~o of patients with acute leukemia in remission were lectin-positive. Contrary to the cells of acute leukemia, only 9 of the CLL patients examined had > 80~o of PNA ÷ cells.

The presence of high levels of PNA ÷ cells ( >_ 15~o) in the peripheral blood of 13 out of 25 (52°,,~) patients with childhood ALL, was reported by Levin et al. [3]. It was further suggested by these authors that a high level of PNA + cells may serve as an indication for poorer prognosis among these patients.

Attempts to find a correlation between the level of PNA ÷ cells and the expression of B or T cell surface markers did not give clear results (Table 2). Thus, the percentage of

TABLE 2. PNA + CELLS IN PERIPHERAL BLOOD LEUKOCYTES OF PATIENTS WITH ACUTE LYMPHOCYTIC LEUKEMIA

Cell type*

Number Number of patients with of different percentages of

patients PNA * cells examined < 2 2-19 20-49 50-79 80-100

T ALL 37 17 5 1 4 10 Pre-B ALL 7 6 0 1 0 0 B ALL 3 l 1 0 0 1 Null ALL 11 6 1 1 1 2

* T ALL, acute lymphocytic leukemia with T cell surface markers; pre-B A L L , acute lymphocytic leukemia with cytoplasmic immunoglobul in expression; B ALL, acute lymphocytic leukemia with surface immunoglobul in ex- pression; null ALL, non-B, non-T acute lymphocytic leuke- mias. Marker studies on this group of patients will be pub- lished separately (in preparation).

Binding of peanut agglutinin to human lymphocytes 125

l e c t i n -pos i t i ve cells a m o n g T A L L pa t i en t s d id no t differ s ign i f ican t ly f r o m the unclass i -

fied A L L .

W e c o n c l u d e tha t whi le the p a t t e r n o f P N A b i n d i n g to n o r m a l h u m a n p e r i p h e r a l

b l o o d l y m p h o c y t e s and to h u m a n t h y m o c y t e s is in l ine wi th the p r o p o s a l tha t the

r e c e p t o r for this lec t in is a m a r k e r for i m m a t u r e cells, this r e l a t i o n ho ld s o n l y pa r t i a l l y

w h e n cells of l e u k e m i c pa t i en t s a re e x a m i n e d . T h e f ind ing tha t a c o n s i d e r a b l y l a rger

p e r c e n t a g e o f pa t i en t s wi th A L L are l ec t in -pos i t ive , t han are pa t i en t s wi th C L L , is in

a g r e e m e n t wi th the n o t i o n tha t cells in the f o r m e r l e u k e m i a s a re m o r e i m m a t u r e than

t hose of the l a t t e r o n e s : m o r e o v e r , as expec ted , u p o n r emis s ion the p e r c e n t a g e o f pos i t i ve

cells decreases . T h e r e is, h o w e v e r , a s igni f icant n u m b e r of A L L cases w h o s e m o n o n u c l e a r

cells do n o t b ind the lect in and , on the o t h e r hand , the re are cells o f C L L pa t i en t s tha t

a re l ec t in -pos i t ive . It w o u l d appea r , the re fore , tha t at least for the t ime being, P N A

b i n d i n g to cells c a n n o t serve as a d i a g n o s t i c a id for the c lass i f ica t ion of l eukemias .

R E F E R E N C E S

1. BALLET J.-J- FELLOUS M., SHARON N, REISNER Y. & AGRAPART M. (1980l Reactivity of human lymphoid and lymphoblastoid cells with peanut agglutinin: detection of a blood cell subset which lacks detectable membrane HLA. Scand. J. Immun. 11, 555.

2. BOYUM A. (19761 Isolation of lymphocytes, granulocytes and macrophages. Scand. J. Immun. 5 (Suppl. 5), 9. 3. LEVIN S.. RUSSELL E. C.. BLANCHARD D.. McWILLIAMS N. B., MAURER H. M. & MUHANAKUMAR T. (1980)

Receptors for peanut agglutinin in childhood acute lymphoblastic leukemia: possible clinical significance. Blood 55, 37.

4. LONDON J.. PERROT J. Y., BERRIH S., LAROCHE L. & NIAUDET P. (1979} Peanut agglutinin. IV. A tool for studying human mononuclear cell differentiation. Stand. J. Immun. 9, 451.

5. NEWMAN R. A.. UHLENBRUCK G., SCHUMACHER K., MIL A. V. & KARDUCK D. (1978) Interaction of peanut agglutinin with human lymphocytes. Binding properties and topology of the receptor site. Z. Immun- Forsch. 154, 451.

6. RABINOVlCH H., UMIEL T.. REISNER Y., SHARON N. & GLOBERSON A. (19791 Characterization of embryonic liver suppressor cells by peanut agglutinin. Cell. Immun. 47, 347.

7. REISNER Y.. LINKER-ISRAEL! M. & SHARON N. (1976) Separation of mouse thymocytes into two subpopula- tions by the use of peanut agglutinin. Cell. Immun. 25, 129.

8. RElSNER Y., ITZICOVlTCH L., MESHORER A. & SHARON N. (1978) Hemopoietic stem cell transplantation using mouse bone marrow and spleen cells fractionated by lectins. Proc. Ham. Acad. Sci. U.S.A. 75, 2933.

9. REISNER Y.. BINIAMINOV M., ROSENTHAL E.. SHARON N. & RAMOT B. (1979) Interaction of peanut agglutinin with normal human lymphocytes and with leukemic cells. Proc. Ham. Acad. Sci. U.S.A. 76, 447.

10. RICHARD Y.. BOUMSELL L., COPPIN H., MISHALL Z., LEMERLE J. & BERNARD A. (1981) Correspondence between lectin-defined and surface antigen-defined cell subpopulations in the human thymus: its variation during ontogeny. J. Immun. 127, 252.

ll. ROSE M. L. & MALCHIODI F. (19811 Binding of peanut lectin to thymic cortex and germinal centers of lymphoid tissue. Immunology 42, 583.

IZ SHARON N. (1980) Cell surface receptors for lectins: markers of murine and human lymphocyte subpopula- tions. In Immunolooy 80 (FoU~EREAU M. & DAUSSET J., Eds.), p. 254. Academic Press, London.