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References Balkwill FR, Oliver RTD (1977) Growth inhibitory effects of Interferon on normal and malignant human haemopoietic cells. Int J Cancer 20: 500-505 - Gutterman J, Yap Y, Buzdar A, Alexanian R, Hersh E, Cabanillas F (1979) Leukocyte Interferon (IF) induced tumor regression in patients (PTS) with breast cancer and B cell neoplasms. AACR 674 - Hill NO, Leob E, Khan A, Pardue A, Hill JM, Aleman C, Dorn G (1980) Phase I human leukocyte interferon trials in leukaemia and cancer. AACR C-167 - Idestrom G, CanteIl K, Killander D, Nilson K, Strander H, Willems J (1979) Interferon therapy in multiple myeloma. Acta Med Scand 105: 149-154 - Mellstedt H, Bjorkholm M, Johansson B, Ahre A, Holm G, Strander H (1979) Interferon therapy in myelomatosis. Lancet I: 245-247 - Merigan TC, Sikora K, Bredden JH, Levy R, Rosenberg S (1978) Preliminary observations on the effect of human leukocyte interferon in non-Hodgkin's lymphoma. N Engl J Med 29:1458-1453 - Osserman EF, Sherman WH (1980) Preliminary results of the American Cancer Society (ACS) - sponsored trial of human leukocyte Interferon (FI) in multiple myeloma (MM). AACR 643 67
Haematology and Blood Transfusion Vol. 26 <strong>Modern</strong> Trends in Human Leukemia IV Edited by Neth, Gallo, Graf, Mannweiler, Winkler © Springer-Verlag Berlin Heidelberg 1981 Methods and Clinical Relevance of Terminal Deoxynucleotidyl Transferase Determination in Leukemic Cells* R. Mertelsmann, M. A. S. Moore and B. Clarkson A. Summary Terminal deoxynucleotidyl transferase (TdT) is a unique DNA polymerase whieh is only found in immature eells of lymphoid lineage (pre-T/pre-B). Beeause of this restricted distribution of TdT, bioehemieal and immunofluoreseenee teehniques have been employed to determine the distribution of TdT phenotypes in human leukemias and lymphomas, showing high levels of TdT in ",95 % of aeute lymphoblastie leukemia (ALL) and lymphoblastie lymphoma (LBL) , ",50% of patients with aeute undifferentiated leukemia (AUL), ",10% of patients with aeute nonlymphoblastic leukemia (ANLL), and ",30% of patients with ehronie myeloid leukemia (CML) and other myeloproliferative (MPS) or myelodysplastie (MDS) syndromes in blast erisis. High levels of TdT aetivity are assoeiated with a c1inieal response to remission indueing therapy with vineristine and prednisone in a high proportion of patients (50%-90%), irrespeetive of clinieal and morphologie diagnosis. Preliminary studies furthermore suggest that TdT might serve as a sensitive indieator of subelinieal disease in ALL in eomplete remission. B. Introduction Terminal deoxynuc1eotidyl transferase is a unique DNA polymerase which adds deoyribonuc1eotides onto an appropriate primer mole- * Supported by Grant Number CA-20194 awarded by the National Cancer Institute, V.S. Dept. of Health, Education, and Welfare, and by the Gar Reichman Foundation eule in the absence of any direeting template polynuc1eotide (Bollum 1979). Under physiologie eonditions TdT is restrieted to thymoeytes and to a subpopulation of bone marrow lymphoeytes whieh exhibit eharaeteristies of immature T (Ineefy et al. 1980; Silverstone et al. 1976) orB eells (Janossy et al. 1979, 1980; Vogler et al. 1978). All other eell types, inc1uding myeloid, erythroid, and putative pluripotent stern eells, do not exhibit deteetable TdT aetivity (Mertelsmann et al. 1979a). Beeause of the restrieted distribution to early developmental stages of lymphoeytes, which loose TdT aetivity during final maturation into cireulating eells, it has been speeulated that TdT might play a role in the generation of immunologie diversity (Bollum 1979). Figure 1 depicts a sehematie model of human hematopoiesis and the eell types to which TdT appears restrieted under physiologie eonditions. Sinee the first report by Kung et al. (1978) about high levels of TdT in ALL, this enzyme has been widely employed in the phenotypic analysis of hemopoietie neoplasias (Bollum 1979; Donion et al. 1977; Gordon et al. 1978; J anossy et al. 1980; Kung et al. 1978; Mertelsmann et al. 1978a,b, 1979a,b, 1981; Modak et al. 1980; Sarin et al. 1976; Vogler et al. 1978). This report reviews eurrent teehnologies for demonstration of TdT and its signifieanee in the c1inical and pathophysiologie evaluation of human leukemias and lymphomas. c. Techniques for Demonstration of TdT Several bioehemical methods have been developed to quantitate TdT activity in human tissues, yielding qualitatively similar results. In order to be able to study routinely obtained 68
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Haematology and Blood Transfusion 2
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Dr. Rolf Neth, Universitäts-Kinder
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Participants of the Meeting Aaronso
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Kurth, R, Friedrich-Miescher-Labora
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Wilsede Scholarship Holders (Grante
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Preface Gut ist eine Lehrart, wo ma
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Personal and scientific discussion
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Acknowledgement We should like to t
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Wilsede, June 21 1978 Memorial Trib
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limiting benign lymphoproliferative
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this most unlikely, however (for re
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longer subject to the same control
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Bregula U, Wiener F, Harris H (1971
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fresh lymph node biopsies from ten
Page 29 and 30: Fig. 3. Binucleate mitosis in an ob
Page 31 and 32: y Fig. 4. Schematic diagram of post
Page 33 and 34: than that among patients with Stage
Page 35 and 36: Tests of binding affinity, agglutin
Page 37 and 38: N Engl J Med 297: 963-968 - Green I
Page 39 and 40: Clinical Aspects
Page 41 and 42: "ring chromosome". "Mar" is marker,
Page 43 and 44: 191 chromosomally abnormal patients
Page 45 and 46: tumors of both African and non-Afri
Page 47 and 48: Haematology and Blood Transfusion V
Page 49 and 50: Retrospective group 26/93 evolved l
Page 53 and 54: Treatment
Page 55 and 56: advantage in terms of duration of f
Page 57 and 58: leukaemia. An analysis of 168 patie
Page 59 and 60: 11. Treatment Program Chemotherapy
Page 61 and 62: efractory congestive heart failure
Page 63 and 64: period (Sequence II) was gene rally
Page 65 and 66: children with a multiple drug proto
Page 67 and 68: DAYS 1 5 10 15 20 25 30 35 , I I I
Page 69 and 70: Biologically Drug Outcome fit" sens
Page 71 and 72: may counterbalance the potential be
Page 73 and 74: Ht (0--0) 50 40 30 20 10 0 Platelet
Page 75 and 76: 10 7 ,-----------------------------
Page 77 and 78: 11. Haematological and Biochemical
Page 79: 0\ 0\ INTERFERON ... 10 3 rl E "'
Page 83 and 84: ("pre-B" phenotype) has been descri
Page 85 and 86: human bone marrow cells exhibit the
Page 87 and 88: 10 9.,0 MBKCC .. .,4 B prot:oool 1'
Page 89 and 90: demann W, Büehner T, Arlin Z, Gee
Page 91 and 92: -...l 00 a-tLDRENS CAt«:ER S TU>Y
Page 93 and 94: prognostic characteristics under in
Page 95 and 96: significantly higher in HR} (14.5%,
Page 97 and 98: normal or elevated immunoglobulin l
Page 99 and 100: References Andreeff M, Darzynkiewic
Page 101 and 102: R GROUP 1 ( 1970 - 1971 ) 17 PTS. R
Page 103 and 104: Table 3. Influence of initial featu
Page 105 and 106: LIJ (f) a.. < -1 LIJ Q: u. 0 >- I-
Page 109 and 110: me nt indicated that additional the
Page 111 and 112: 42:2123-2134 - Chessells JM, Cornbl
Page 113 and 114: Baum et al. 1979). This study was b
Page 115 and 116: 1. 0 '--'1--" 0.9 e: o .- VI VI E C
Page 117 and 118: e: o e: 1. 0 ,.--...-- 1'"'\ ......
Page 119 and 120: In conclusion, we can state that th
Page 121 and 122: Haematology and Blood Transfnsion V
Page 123 and 124: INDUCTION VCR + PRED + ASNASE (wk 3
Page 125 and 126: SCHED.INT 1 MTX • ! Adria MTX •
Page 127 and 128: ~ looh---------ooooc>----o---c>----
Page 129 and 130: Consoll- Irra .tlon t) .tlon Indu
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Highrisk Standard risk Table 1. Cli
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Cumulo/ive comp/el9 remission 1.0 .
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10° 8 = 6 ö 4 Non T Cell .~ :ö
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Haematology and Blood Transfusion V
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% SURVIVAL 100 ~ ~II 0 80 60 40 - -
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a) Percentage of donor cell mitoses
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Table 5. Results of bone marrow tra
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...... w N Table 7. Clinical result
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agent had to be added and that in t
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C. Chronic Myelogenous Leukemia The
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Patients with a suitable donor rece
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No. Recurrent Leukaemia Other death
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followed by dialysis against isoton
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HS, HL- Heme >_ 40 0 HS I ;:. =====
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knowing at present. These genes may
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esistance to antifolates. In Vitro
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a DNA probe specific for the gene t
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translocation between chromosomes 9
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Table 1. Subdivision of 1827 Myelo-
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NORMAL CELL ®
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some change of the primary type, in
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F. Transformation of Mouse Bone Mar
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were carried out with 3H-MTX in the
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the lysozyme cDNA prepared from ovi
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et al. 1976). The BJAB cellline in
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dalton bands corresponding to light
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1979). The EBV-carrying cell lines
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producer lines (DAUDI and P3HR-1),
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G, Giovanella B, Westman A, Stehlin
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Dc;bt CT + i J.I9 Raji; 1. i .. 100
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c. Discussion During infectious mon
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C. Nonepisomal Viral DNA We have us
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A 260 3.0 A B 1670 70 N2 2.0 c: E .
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vivo. Nature 260:302-306 - Kirk JTO
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Fig. 1. a Polyacrylamide gelelectro
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1979). The lymphocyte subpopulation
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lysin 0 antigens are found, and mix
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Pope JH (1978) Long-term T cell-med
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Haematology and Blood Transfnsion V
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Charaeteristie Morphologie maturati
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after TPA-induction (Table 3). The
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PA (1978) Reactivity of a rabbit an
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Blood Monocyte. B lood Monocytes 1-
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and nonhistone proteins can serve a
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The findings of the evaluation of s
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Fig. 3. Tumor grawth of L 428 cells
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Cell biological and Immunological A
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consideration. Sanel (1973) obtaine
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endotoxin serum, were used. A parti
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monocytic leukemia cell line in cul
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B. Isolation, Characterization and
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don al hemopathies generally displa
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nitors. Blood Cells 6:595-607 - Min
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\I) ....I ....I IU Jl 200 r 100 90
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their haemopoietic cells' ability t
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normal normal ALL ALL ALL persons p
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64.5 { V)
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References Biermann E, Neth R, Gros
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ehambers, the growth pattern observ
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References Anderssen LC, 10kinen M,
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REH CELL UNE TO CALLb~Ö ~q~ V.M. c
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the macrophage series in which the
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Table 1. Production of factor depen
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Table 1. Characteristics of the adh
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Fig. 3. A human marrow culture at 6
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300 250 • HYPERPROLIFERATIVE PATT
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Fig. 9. Nonadherent population from
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The cobblestone areas were often no
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L (1976) Induction of colony format
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B. Materials and Methods J. Tissue
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Fig. 2. Ultrastructural appearance
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Table 1. Monoclonal antibody reacti
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al. 1979; Janossy et al. 1979) is e
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to which this is an exact replica o
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inger JL (1976) Distribution of la-
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I region antigens were found to be
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have been reported to be present on
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V. FunctionalAssays Assays for PHA
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Functional studies of UCHT1 separat
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40 ,...... (5 L.. C o u 30 .9 ~ .~
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with immunohistochemieal methods (L
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can be dissected by cloning. Utiliz
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Table 1. Origin and marker profile
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complete identity of gp 100 from no
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Table 1. Reaction of single anti se
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media. Following 24-h incubation, c
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nonleukemic eells (Greaves 1979). L
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Fig. 2. Cytocentrifuged smears of b
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Modification of amino-groups of hum
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disturb this balance in the directi
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The majority of human blood lymphoc
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of the EBV infected B cells. Howeve
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and immunoglobulins. J Immunol 120:
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" K 10 • j ~ ALL RNlL AL wtfh les
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tion akuter Leukämiezellen in "spo
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D. Results and Discussion J. Acute
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selectively responsive to different
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l1J ~ 100 ~ Q. ::) l1J Z 0 ~ z >- .
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Table 2. Growth inhibition of S49 e
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Reverse Infectious ASC/2X1Q6 transc
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munized animals to 1316 tumor cells
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Haematology and Blood Transfnsion V
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Fig. 3. Expression of retroviral gp
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pg70. J Exp Med 143: 151-166 - McGr
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Table 1. Expression of tumor antige
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Virological and Molecularbiological
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Table 1. Oncogenic properties of re
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also Fig. 1). It followed that the
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whether this sequence is related to
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zed in infected cells argue that th
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a specific viral transforming prote
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detect viral RNA as the only charac
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Eisen H (1980) Myeloproliferate vir
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p,60- -34K - 1 2 3 4 Fig. 1. In vit
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.. p - • a'T ... I .t'l Fig. 4. D
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Fig. 2. Photomicrographs of NY68 in
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DEAE (o(lJmn 'liI GI -2 2 o e s 1
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vity was deterrnined. For endogenou
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pr 180 - - - P 12/15 - A B c D E F
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85,- 66- -27 - -66 40- 27- -1'9 19-
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= 0.5 o E Q. o ILI 0.4 (J) « ILI .
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Table 1. Oncogenic potential of avi
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100 III GI c: o "8 80 > w
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Expt. No. Infecting virus Proportio
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Haematology and Blood 'fransfusion
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sequences, provided its gag portion
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77:3009-3013 - Hu SSF, Moscoviei C,
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RI 2.2 md and Hind III 2.6 md leuke
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RELATIONSHIP OF ENDOGENOUS AND EXOG
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E ........ E Cl. U E :::l .... Q) (
Page 447 and 448:
References Astrin S (1978) Endogeno
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the same animal were examined (e.g.
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I) 'IQ a \0 - CL -- Fig. 2. Restrie
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~ ..................... ~ Cell 3' 5
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a :Sa,C! + . ~ 111 i' ( " '~ .J. b
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Table 1 Sampie Tissue TS fragment"
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a b Sac I / rep ._-_.-_ K J nl rep
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Haematology and Blood 'fiansfusion
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Osteopetrosis and osteosarcomas occ
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Lymphoma cellline Table 1. T and B
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Table 2. In vivo growth and oneogen
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Table 1. Transforming aetivity of c
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Table 3. Transformation aetivity of
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- Shoemaker C, Goff S, Gilboa E, Pa
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indieate a shifting of target eell
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Table 2. Expression of Jl and Sourc
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~ i ,..1---------,,':::::::::::::::
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fragment of pBR322 DNA, a 3.0-kbp S
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Mak TW (1979) Proc Natl Acad Sci US
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Table 1. Transformation of 3T3 cell
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inserts in the genome of rapidly tr
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pulations, it seemed likely that th
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contains antigens which react with
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le of reacting in sensitive radioim
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gic approach. In: Hiatt HH, Watson
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12 Eco R [ Fig. 1. Hybridization pa
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10 - '1~ )( -~ S:! E A U I o 1 2
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Haematology and Blood 'fransfusion
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cultures. These cells did not conta
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indication of malignant T-cells in
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Fig. 3. Thin-section electron micro
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Table 3. Lack of detectable related
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J. HTLV Was Present in the Primary
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specific signals into nonspecific s
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Table 2. Distribution of HTLV-relat
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u 10 ~ 20 ~ :c 30 a 40 o Fig. 1. Ki
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SSV TrS-gp labeled effectively with
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60 HFF/SSV cDNA CELL RNAs: 0 HF/SSV
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Antisera a Table 1. Immunofluoresce
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Haematology and Blood 'ftansfusion
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y competition RIA yielded virtually
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SiS'V' gp70: ANTIGEN, ANTI BODY, IM
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type-C virus p30 antigen in cells f
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c. Results The fractionated whole-b
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induced antibodies as well as exper
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- Fig. la-f. Retravirus particles p
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eplication of infecting murine leuk
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immunodeficiency, multic10nal lymph
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Bovine leukemia --, proteins of 499
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Human T- cell - -, characterization
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RNA -, of tumor virus 439 Rous sarc
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Haematology andBlood Transfusion Su
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