Low dose arabinosyl cytosine for treatment of myelodysplastic syndromes and subacute myeloid leukemia

LeukemiaResearchVol.7, No. 4, pp. 539-545, 1983.

0145-2126/83 $3.00 + 0.00 © 1983PergamonPressLtd.

Printedin GreatBritain.

LOW DOSE ARABINOSYL CYTOSINE FOR TREATMENT OF MYELODYSPLASTIC SYNDROMES AND SUBACUTE MYELOID LEUKEMIA MICHELE BACCARANI, ALFONSO ZACCARIA, GIUSEPPE BANDINI, G1OVANNA CAVAZZINI, RENATO FAN1N and SANTE TURA Istituto di Emal;ologia "Lorenzo e Ariosto Ser~tgnoli", Universita di Bologna, e Cattedra di Ematologia, Universita di Trieste, ltalia

(Received 9 March 1983. Accepted 12 April 1983) Abstract--Several agents, including arabinosyl cytosine (ARA-C) at a low concentration, can induce leukemic myeloblasts to mature to a variable extent. The therapeutic implications of this observation are worth investigating. A few case-reports have shown that low dose ARA-C can be useful for treatment of the myelodysplastic syndromes (MDS) and of acute myeloid leukemia (AML). However, no information is available yet on the proportion of patients who can be expected to respond. We treated by low dose ARA-C (20-30 mg/sqm/day i.v. or i.m. for 7-10 days) 20 consecutive patients. A complete remission of 5 months was obtained in one of nine cases of subacute myeloid leukemia (SAML). A partial remission (complete normalization of blood counts with a slight excess of marrow blast cells) was obtained twice in one of 11 cases of MDS. An increase of Hb level (more than 11.5 g/dl) was obtained and maintained for 12 months in a case of MDS. A short-lasting increase of granulocyte count was obtained in another two cases of MDS and SAML respectively. It is suggested that low dose ARA-C can advantageously modify the proliferation to maturation imbalance of leukemic cells by slowing down cell proliferation rate. However, the proportion of patients who respond is probably low. This treatment is at a very early experimental stage and should be probably limited to selected cases of MDS and subacute or acute myeloid leukemia.

Key words: Arabinosyt cytosine, myeiodysplastic syndrome, myeloid leukemia, cell differentiation.

INTRODUCTION THE DEFINITION of hemopoietic dysplasia or myelodysplastic syndrome (MDS) covers a broad spectrum of poorly defined hematologic diseases of unknown etiology, which are commonly regarded as preleukemic states [6, 15, 16, 23]. In some instances the MDS cannot be easily distinguished from myeloid leukemia, either acute or subacute [5, 6, 15, 16, 23] and its relationship to leukemia is confirmed by a number of laboratory investigations and by clinical studies showing evolution from MDS to overt leukemia [15, 16, 22, 231. In acute myeloid leukemia (AML), blast cell proliferation leads to a large accumulation of leukemic cells in the body. At the same time, leukemic cell maturation is so defective as to reduce the production of mature blood cells down to levels that are incompatible with life [18, 19]. In some cases of leukemia, blast cell accumulation is slower and cell maturation is less defective, so that clinical features are less dramatic. These cases have been defined as subacute or smoldering leukemia [5, 15, 16, 21]. In the MDS proliferation and

Abbreviations: AML, acute myeloid leukemia; ARA-C, arabinosyl cytosine; CMML, chronic mydomonocytic leukemia; MDS, myelodysplastic syndrome; PMN, polymorphonuclear, neutrophilic; RA, refractory anemia; RAEB, refractory anemia with excess of blast cells; RAEB-T, refractory anemia with excess of blast ceils, in transformation; SAML, subacute myeloid leukemia. Correspondence to: Dr. Michele Baccarani, Istituto di Ematologia "L.e.A. Ser~tgnoli", Ospedale S. Orsola, 4013 8 Bologna, Italy. 539

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MICHELEBACCARANIet al.

m a t u r a t i o n d e f e c t s a r e u s u a l l y less e v i d e n t a n d t h e c l i n i c a l c o u r s e is m o r e b e n i g n [15, 16]. O n t h a t a c c o u n t , w h i l e t h e p r e s e n t t r e a t m e n t o f A M L is b a s e d o n c y t o t o x i c a g e n t s , a i m i n g at eradicating leukemia, the management of MDS and of subacute myeloid leukemia ( S A M L ) is o f t e n b a s e d o n s u p p o r t i v e t h e r a p y a n d b l o o d t r a n s f u s i o n [12, 15, 16, 22, 23]. T h i s c o n s e r v a t i v e p o l i c y o f t r e a t m e n t c a n b e j u s t i f i e d b y the r e l a t i v e l y l o n g " s p o n t a n e o u s " d u r a t i o n o f M D S a n d S A M L , b y t h e r e l a t i v e l y o l d a g e o f m a n y p a t i e n t s a n d by the observation that too many such patients fail to achieve a remission with intensive cytot o x i c t h e r a p y , as t h e y die d u r i n g t h e r a p y - i n d u c e d b o n e m a r r o w a p l a s i a [1, 12, 15, 16, 22, 23]. T h e r e is s o m e e v i d e n c e t h a t s e v e r a l a g e n t s , i n c l u d i n g classic a n t i l e u k e m i c d r u g s a t a l o w concentration, can advantageously modify the proliferation to maturation imbalance of l e u k e m i c cells a n d c a n i n d u c e t h e m t o m a t u r e [I0, 24]. I n 1979, w e r e p o r t e d o n t h e succ e s s f u l t r e a t m e n t o f a c a s e o f M D S b y l o w d o s e a r a b i n o s y l c y t o s i n e ( A R A - C ) [4]. T h i s o b s e r v a t i o n was t a k e n u p b y o t h e r s a n d w a s c o n f i r m e d in a f e w o t h e r c a s e s o f A M L [13, 20]. W e r e p o r t h e r e o n t h e e f f e c t o f l o w d o s e A R A - C in 20 c o n s e c u t i v e p a t i e n t s a f f e c t e d by MDS and SAML. PATIENTS

AND METHODS

Twenty patients with MDS and SAML were entered into the study. Morphological examination of Giemsastained marrow and blood cells, several cytochemical reactions (peroxidase, naphthol-ASD-esterase with and without prior NaF incubation, ~-naphthyl esterase, chloroacetate esterase, PAS and Pearl's reaction for iron) and serum lysozyme concentration, were used for classification, according to the suggestions of the FrenchAmerican-British group [5, 6]. All patients had been seen and controlled for several months (2-36 months) prior to ARA-C treatment. At the time of first ARA-C treatment, seven cases were classified (Table 1) as refractory anemia with excess of marrow blast cells (RAEB), four cases as RAEB in "transformation" (RAEB-T), one case as chronic myelomonocytic leukemia (CMML) and eight cases as myeloid leukemia, M2 or M4 subtype. These

TABLE 1. CLASSIFICATIONANDFOLLOW-UPOF 20 PATIENTSTREATEDBYLOWDOSEARA-C Classification Case

1 -VF 2--CV 3 - - FI 4 - - CL 5 - - PN 6 - - MG 7 - - SG 8 - - RM 9 - - BI 10 - - MA ! 1 - - MD 12 - - VF 13 - - MF 14 - - VT 15 - - DSF 16 - - MC 17 - - ZE 18 - - FP 19 - - BE 20 - - MU

Survival* (months)

Age

Sex

At first diagnosis

At treatment

At last follow-up

From diagnosis

From treatment

24 26 45 73 28 70 57 28 38 42 57 57 40 55 63 61 52 60 69 77

M M M M M M M M F M M F F M M M M F F M

RA RA RAEB RA RAEB RAEB RAEB RAEB-T RAEB-T RA RAEB-T RAEB-T RAEB-T SAML (M2) RAEB SAML (M4) SAML (M4) SAML (M4) RAEB CMML

RAEB RAEB RAEB RAEB RAEB RAEB RAEB RAEB-T RAEB-T RAEB-T RAEB-T SAML (M2) SAML 0vl2) SAML (M2) SAML (M2) SAML (M4) SAML (M4) SAML (M4) SAML (M4) CMM L

RAEB RAEB AML (M2) RAEB AML (M2) RAEB AML (M2) AML (M2) AML (M4) RAEB-T AML (M2) AML (M2) AML (M2) SAML (M2) AML (M2) SAML (M4) SAML (M4) SAML (M4) AML (M4) CMML

62 + 39+ 19 20 11 27 + 31 14 22 46 6 9+ 9 35 9 8+ 6 14 + 15 36

26 + 13+ 14 8 3 2+ 22 7 18 42 4 3+ 3 30 3 4+ 4 9+ 3 40

* + Indicates that the patient is still alive.

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541

cases were defined as "subacute" because of the slow rate o f progression of the disease, but on pure morphological grounds at marrow examination, they met the criteria for the definition of acute leukemia [5, 6]. On subsequent follow-up, 10 patients developed overt AML in less than 1 year (Table 1). Patient No. 17 had received total nodal irradiation and MOPP chemotherapy for Hodgkin's disease 4 years before developing leukemia. The case of patient No. 7 was previously reported elsewhere [4]. Since first diagnosis, all patients had received regular blood transfusion and all cases of RAEB had failed to respond to vitamins B6 and BI2 and to folinic acid. Eight patients had been occasionally given corticosteroids at a low dose, without any effect. Four patients had failed to respond to oxymetholone. No patient had received cytotoxic drugs. At the time of study, all patients were anemic (Hb concentration ranging between 4.7 and 11.7 g/di) with a reticulocyte count lower than 50 × 109/1, 16/20 patients had a platelet count lower than 100 × 109/1 and 12/20 patients had an absolute neutrophilic polymorphonuclear (PMN) count lower than 1.5 × 109/1. Peripheral blood blast cells were identifiable in 11 of 20 cases (range 2-76%, or 0. i =7.4 × 109/1 ). The marrow was hypercellular in all cases except No. 1 and 2, who had a hypoplastic marrow with several pockets of ineffective hemopoiesis. Spleen was palpable in patients No. 3, 9 and 16, 3-5 cm below costal margin. Lymphadenomegaly, gum hypertrophy and skin involvement were found in no case. Blood counts and differentials were performed prior to and at 3-5-day intervals during and after treatment. Bone marrow examination was repeated at least once, 1-2 weeks after treatment. Chromosome studies were performed in nine cases. Treatment consisted in 7-10-day courses of ARA-C, 20-30 mg/sqm/day. In 11 patients, ARA-C was given as a single i.e. or i.m. injection every 24 h. In the other nine patients, a 12 h schedule was used. Thirteen patients had a single course of treatment, four patients had three courses, two patients had four courses, and one patient (case No. 18) had seven consecutive courses, at 4-week intervals.

RESULTS The clinically relevant results of treatment by low dose ARA-C are shown in Table 2. A complete marrow and blood remission was obtained only in one patient (case No. 18) who received the first course of ARA-C (25 mg/sqm/day i.v., in two divided doses, for 7 days) 5 months after the diagnosis of SAML(M4). Leukemic blast cells disappeared from peripheral blood, platelet count increased up to more than 100 x 109/1 and bone marrow became slightly hypoccllular, with 3507o blast cells (Fig. 1). After a second course of ARA-C, a complete marrow and blood remission was obtained and it was maintained with 5-day monthly treatments of ARA-C, at the same dose and schedule. Leukemia relapsed after 5 months, but the patient remained on the same treatment and she was alive and well after 5 more months.

TABLE2. RESULTSOFTREATMENT

Complete remission

: 1 case (No. 18, SAML-M4)

Partial remission

: 1 case (No. 7, RAEB)

Isolated increase of Hb level

: 1 case (No. 9, RAEB-T)

Isolated increase of granulocyte count : 2 cases (No. 2, RAEB; No. 17, SAML-M4)

A complete normalization of blood counts and a partial marrow remission (decrease of blast cell proportion to less than 1007o)was obtained twice in a patient with RAEB (case No. 7) with two courses of ARA-C (30 mg/sqm/day i.v. for 7 days). This case was reported in detail elsewhere [4]. The first remission lasted 5 months and the second 3 months. A third and a fourth course of treatment were unsuccessful and the patient died of AML(M2). A significant increase of Hb concentration (more than 11.5 g/dl) was obtained and maintained for 12 months in a 38-year-old lady with RAEB-T (case No. 9), with four

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FIG. 1. Effect of repeated courses of low-dose arabinosyl cytosine (ARA-C) in patient No. 18, with subacute myeloid leukemia, M4 subtype. A complete blood and marrow remission was obtained after the 2nd course of treatment and was maintained for 5 months. ARA-C was given at a dose of 25 rag/reX/day i.v., in two divided doses, for 7 days.

7-day courses of ARA-C (30 mg/sqm/day i.v.). Prior to treatment, the Hb level was maintained at 10 g/dl with weekly red cell transfusions. In this case, the treatment corrected dyserythropoiesis and decreased slightly marrow cellularity, but did not modify bone marrow blast cell concentration and did not increase PMN and platelet count. In another two cases (No. 2 with RAEB and No. 17 with SAML-M4 developing after combined radiochemotherapy for Hodgkin's disease), a significant (more than 2 x 109/1) but short-lasting (40 and 50 days respectively) increase of blood PMN was observed, together with a significant decrease of bone marrow blast cells (to less than 50°7o of pretreatment value), but without any measurable effect on Hb level and on platelet count. Marrow cellularity remained unchanged in both cases. In the remaining 15 patients, treatment by low dose ARA-C failed to induce any clinical benefit, although a significant decrease of blood blast cells (to less than 107o) and of marrow blast cells (to less than 50070 of pretreatment estimate) was shown in three and five cases respectively, together with a significant decrease of marrow cellularity. In this study, karyotype constitution of marrow cells was determined in nine cases only. Three patients (cases No. 4 and 6 with RAEB and case No. 18 with SAML-M4) had a normal karyotype. Two of them failed, while the third one got a complete remission. Five patients (cases No. 1 and 5 with RAEB, cases 8 with RAEB-T, cases No. 12 and 13 with SAML-M2) had a mixture of normal and abnormal metaphases. All of them failed. One patient (case No. 17 with SAML-M4 developing after radiochemotherapy for Hodgkin's disease) showed only abnormal metaphases, with a 44 X Y - - 5 , - - 7 , - - 15,-- 15, + M 1, + M2 clone. Upon treatment by low dose ARA-C, a significant increase of blood PMN together with a significant decrease of marrow blast cells was observed.

Low dose ARA-C

543

DISCUSSION The proliferation and maturation defects of leukemic cells are not absolute [18, 19]. In vivo, the rate of proliferation can vary much from one case to another and in the same case and a partial maturation of leukemic cells is usually detectable. As a matter of fact, morphological and cell-marker based classification of leukemia depends on the residual maturation ability of leukemic blast cells. In vitro, there is some evidence that a number of agents, including some classic cytotoxic drugs, can modify the proliferation to maturation imbalance of leukemic cells and can induce them to mature to a variable extent [7, 10, 14, 17, 24]. The potential for treatment of these observations is iinknown. A few case reports suggested that such a potential is worth investigating. Upon administration of ARA-C at a low dose (20-30 mg/sqm/day) a partial remission was obtained twice in a case of RAEB [4] and a complete remission was obtained in five cases of subacute or acute myeloid leukemia [13, 20]. These observations raise two main questions, (1) what is the actual proportion of patients who can benefit from such a treatment and (2) what is the actual mechanism of action of low dose ARA-C. This paper provides data relevant to the first question. It was found that a partial remission was obtained twice in one of 11 cases of MDS and a complete remission in one of nine cases of a SAML, for a response rate of 10%. In another three cases, an isolated increase of Hb concentration or of PMN count was observed. The increase of Hb concentration was of clinical benefit, as it was maintained for 12 months and eliminated the need, which had been previously established, for frequent red cell transfusion. The actual 10% minimum response rate figure should be taken only as an indicator and this confirms the potential usefulness of treatment by low dose ARA-C, but suggests that only a minority of the patients can be expected to benefit from it, at least with the dose and schedule that were used in this study. As a matter of fact, in the five AML patients who were successfully treated by Moloney and Rosenthal [20] and by Housset and coworkers [13], ARA-C was given for a significantly longer period (17-25 days). By contrast, a large cooperative study of RAEB failed to show any benefit from shorter courses of low dose ARA-C (20 mg/sqm/12 h x 4, monthly) [9]. The proportion of patients who responded to treatment was too low to look for a correlation with disease subtype and features or with karyotype constitution. Information on many more patients is needed. The mechanism of action of ARA-C at a .low dose is unclear. At higher doses (more than 100 mg/sqm/day) ARA-C is known to inhibit DNA-synthesis after transformation to ARA-CTP [8, 25]. Hence, inhibition of cell replication, cell accumulation in S-phase, and cell death follow [2, 3, 8, 28] so that the usual effect of a standard dose of ARA-C is marrow hypoplasia or aplasia [12]. However, there is evidence that the mechanism of action of the drug can be more complex, as ARA-C is incorporated by leukemic cells also outside S-phase (e.g. in G1 phase) and can inhibit also protein synthesis [25, 26, 27]. At a very low concentration, ARA-C could slow down cell proliferation rate, so as to allow the cells to display their whole residual maturation potential. In that case, "maturation" would be a consequence of the inhibition of proliferation, with a variable degree of cell death. On the other hand, the effect of ARA-C on the synthesis of DNA and/or other macromolecules could result in a "primary" maturation effect, without a significant or relevant inhibition of cell proliferation. We favour the former hypothesis and some investigations have provided evidence that the induction of maturation is associated to the inhibition of proliferation. In a clonal assay, Chang and McCuUoch [7] showed that 12-Otetradecanoyl phorbol acetate (TPA) promoted ANA-esterase activity of leukemic cells only at proliferation-inhibiting concentrations. Similar results had been obtained by Huberman and Callahan [14] with the human promyelocytic leukemia cell line HL-60.

544.

MICHELE BACCARANIet al.

Krystosek and Sachs [17] found that ARA-C induced lysozyme production in a mouse myeloid leukemia cell clone at a concentration (0.1 ~g/ml) that caused 85°70 inhibition of growth. The data reported in this paper cannot help much to clarifY, that point. However, a significant reduction of marrow cellularity was observed in three of five cases who responded and in another five cases treatment induced a significant (more than 50070) decrease of marrow cellularity and of marrow blast cells, without any evidence of maturation. A third and plainer explanation should not be overlooked. The effect of low dose ARA-C in some patients could rest on a lower rate of degradation of the drug and/or on a higher sensitivity of some leukemic cell clones, leading to the progressive reduction of leukemic cell number via a classic cytotoxic mechanism. This would allow marrow replenishment by a normal or a "leukemia min" cell population [19]. In conclusion, there is evidence that standard cytotoxic treatment of AML, coupled with high standard supportive care, can be of benefit also in the elderly and can induce a complete remission also in cases developing after a MDS or after radiochemotherapy for other malignancies [1, 11, 12]. Therefore, treatment by low dose ARA-C will remain so far at a very early experimental stage, and would probably be better reserved to the MDS and to selected cases of subacute or acute myeloid leukemia. Its appeal for the clinician rests on a very low cytotoxic cost, which is important in elderly patients, and for the scientist on the opportunity of gaining further knowledge on the biology of leukemic cells and of manipulating their behaviour. Acknowledgements--Work supported by CNR, Finalized Project for Control of Tumor Growth, Contracts No. 82.00222.96 and 82.00428.96.

Note f r o m the Editors: Housset et ai. [13] reported more encouraging results with low dose ARA-C than the present paper. Though the patients material may be different, it should be noted that Housset et al. administered ARA-C subcutaneously every 12 h for 25 or more days, thus probably achieving a rather constant in vivo concentration of the drug.

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