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Prognostic factors in acute myeloid leukaemia
Best Practice & Research Clinical Haematology Vol. 14, No. 1, pp. 65±75, 2001
doi:10.1053/beha.2000.0116, available online at http://www.idealibrary.com on
4 Prognostic factors in acute myeloid leukaemia Bob LoÈwenberg
MD, PhD
Professor of Haematology Erasmus University and University Hospital Rotterdam, P.O. Box 2040, 3000 CA Rotterdam, The Netherlands
The prognosis between subgroups of patients with acute myeloid leukaemia (AML) may range considerably. Haematological, genetic and clinical analysis has provided possibilities for de®ning the heterogeneity of prognosis. This furnishes clinically relevant insights into the probability of treatment response and survival in individual cases of AML and it provides a lead in treatment management. Key words: prognosis; prognostic factors; acute myeloid leukaemia; cytogenetics; ¯t-3 receptor; ¯t-3 mutations; molecular genetics.
INTRODUCTION The group of diseases collectively designated as acute myeloid leukaemia (AML) represents a broad variety of distinct entities. AML arises from a malignantly transformed haematopoietic stem cell or progenitor cell. The cell stage (more or less primitive, multipotent or committed) at which transformation occurs and the residual capabilities of maturation of these cells, may contribute to the phenotypic diversity of this disease.1±3 These variations may be re¯ected in the dierent cytomorphology of the leukaemic blasts, the dysplasia of the other haematopoietic cell lineages and the characteristic sites of presentation of leukaemia (e.g. skin, gingiva etc). The molecular abnormalities leading to full leukaemic transformation vary greatly. Dierences in the molecular pathogenesis may also dictate variation in clinical presentation as well as variation of response to therapy. The understanding that the term AML lumps together a variety of distinct pathogenic conditions, has led to a general appreciation of the importance of delineating AML disease entities more sharply, e.g. AML with t(8;21) or AML1/eight-twenty-one (ETO) fusion gene etc. Each of the genetically de®ned AML diseases carries a typical prognosis and one may anticipate that in the long run these conditions will require speci®c therapy. Acute promyelocytic leukaemia with t(15;17) and the promyelocytic leukaemia gene/retinoic acid receptor alpha gene (PML/RAR-a) fusion gene, treated with all-trans retinoic acid (ATRA) is a classic example of this approach.4 Meanwhile a thorough understanding of the individual prognostic factors has become of signi®cant importance for treatment choice in today's clinical practice. Should the patient be treated with an allograft from an HLA-identical sibling or should 1521±6926/01/01006511 $35.00/00
c 2001 Harcourt Publishers Ltd. *
66 B. LoÈwenberg
the transplant be withheld and reserved in case of relapse? Who should receive an allogeneic HLA matched transplant from an unrelated donar and when? Who should receive high-dose cytarabine as part of their chemotherapy? Who should be entered onto experimental protocols? In addition to cytogenetics in recent years new leukaemia-related prognostic factors (e.g. ¯t-3 mutations) have been recognized, which may have an important impact on prognosis. Here we will review the current knowledge of factors that are predictive of the response to therapy and outcome in adults with AML (Table 1). AGE AND GENDER Age has a strong prognostic impact on the outcome of AML. There is a gradual but consistent decline in the prognosis for patients with AML with increasing age. This negative relationship is continuous from infant to 85 years of age. The latter relationship is apparent as a progressive decrease in treatment response and survival as a function of decade of age (Oxford Collaborative AML Intergroup, pers. comm.). For practical reasons of protocol design, a cut-o point of 60 years is usually used to distinguish older patients. Young and middle-aged adults, i.e. those less than 60 years old, have response rates to remission induction chemotherapy of approximately 75% and approximately 35±40% of them survive following diagnosis for 5 years or more.5±11 In contrast, patients with AML who are more than 60 years old have an overall response probability to induction therapy of 45±55%, and fewer than 10% of these survive for a minimum of 5 years.12,13 While the results of treatment have improved steadily over the last 20 years in younger adults, due to more intensive cytotoxic treatment, the use of stem cell grafts and improvements in supportive care, no signi®cant change in outcome has been noted among individuals of 60 years of age. The outcome for older subjects may even be worse than these estimates would indicate, since many older patients are not referred to hospitals for treatment, and are not registered or evaluated for outcome. Thus these outcome data for aged individuals probably represent overestimations. Why is outcome so unsatisfactory in the aged? The reasons for the poorer outcome of patients of higher age most probably relate to the increased frequency of unfavourable cytogenetics among older patients with AML, a greater frequency of antecedent myelodysplasia, as well as a greater frequency of drug resistance phenotypes. Also, because of the reduced general health of older patients, they cannot withstand intensive chemotherapy as well as younger individuals do.13,14 Several large databases of phase III studies in untreated patients with AML would also suggest that female sex is an independent favourable prognostic parameter for disease-free survival, although of moderate impact only. The intergroup analysis of the Oxford Collaborative AML Intergroup (pers. comm.) has con®rmed that disease-free survival of male patients is somewhat less than that of females. CYTOGENETICS AND MOLECULAR GENETICS Cytogenetic abnormalities are seen in approximately 60% of cases in AML and are highly predictive of response to therapy as well as the probability of relapse.15±18 The translocations t(8;21), inv16 and t(15;17) generally carry a relatively favourable prognosis and these are more commonly seen among younger patients with AML. The fusion genes of each of these translocations have been identi®ed and can be detected
Prognostic factors in acute myeloid leukaemia 67
using molecular probes in reverse transcriptase-polymerase chain reactions (RT-PCR). Among patients with these chromosomal abnormalities or the corresponding molecular abnormalities (AML1-ETO, CBFb-MYH11 (core binding factor beta genesmooth muscle myosin heavy chain gene), PML-RARa) the response to induction therapy is 80% or greater and for those entering remission the probability of relapse is 30±40% resulting in 5 years survival rates of 60±70%. The latter molecular or cytogenetic subsets of AML correlate with certain cytomorphological categories. For instance, t(15;17) or PML/RAR correlates with the French±American±British (FAB) group subtype M3, and inv16 with cytological FAB subtype M4 with eosinophils. However, the cytogenetics and molecular genetics rather than the cytological FAB classes, determine the overriding prognostic signi®cance. In contrast patients with t(6;9), 11q23 (MLL-gene) abnormalities19, monosomies of chromosomes 7 (-7) or 5 (-7) and complex cytogenetic abnormalities (three dierent cytogenetic aberrations or more) generally have a distinctively poor prognosis. In these individuals (when 60 years or younger), the average complete response to induce treatment is 60% with a majority (approximately 80%) relapsing within 2 years so that their survival at 5 years is approximately 15% only.11,20 The cytogenetic and molecular indicators have led to the de®nition of prognostic groups, i.e. favourable risk (with favourable molecular genetics or cytogenetics), unfavourable risk (those with poor-risk cytogenetics) and intermediate risk (all others). Usually, these risk classi®cations are re®ned by considering selected additional prognostic determinants that enhance the value of these cytogenetic or molecular distinctions. The risk-scoring systems that are based upon multiple prognostic factors may permit a more precise approximation of prognosis in individual cases. To keep this practically simple, most collaborative groups have restricted the use of these additional parameters, and only taken into account a limited number of covariables that have a considerable impact. As an example, the Dutch HOVON (Haemato-Oncology Collaborative Group for Adults±Netherlands) group has required a white blood cell (WBC) count of less than 20 109/l in addition to favourable t(8;21) cytogenetics for the good risk category assignment (risk of relapse is 24% at 5 years: Figure 1). The Medical Research Council (MRC) has included the rapid versus late attainment of complete response (i.e. after cycle 1 versus after cycle 2) in their risk score in order to enhance the separation between good risk and poor risk.20
IMMUNOPHENOTYPING Cell stage and cell lineage speci®c markers have been used for immunophenotyping of AML. While the cytogenetic and molecular genetic markers carry a strong prognostic impact, the value of immunophenotyping for the assessment of prognosis has not been ®rmly established. This is true for the myeloid markers CD13, CD14, CD15, CD11b, HLA-DR, CD117 (c-kit) as well as CD34 for which controversial results regarding the prognostic value for response have been obtained in a variety of studies.21 The choice of antibodies, the threshold values for positivity (e.g. percentage positive cells) and clinical variations may explain some of the discrepancy between dierent studies. More importantly, most positive correlations between immunophenotype and prognosis do not hold up in multivariate analysis indicating that other associated determinants exert a greater eect on prognosis.
0
25
50
75
100
0
0 39 4
34 4
27 3
20 2
23 2
Months 60
WBC>20
WBC<=20
24 3
Months 60
WBC>20
WBC<=20
0 96 79
0 297 206
N O WBC<=20 686 422 WBC>20 570 344 Log rank P = 0.27
152 100
(D) No favourable cytogenetics
At risk: WBC<=20 686 WBC>20 570
0
25
50
75
100
182 147
N O WBC<=20 546 269 WBC>20 450 203 Log rank P = 0.82
At risk: WBC<=20 546 WBC>20 450
0
25
50
75
100
(B) No favourable ctyogenetics
91 71
65 58
56 49
Months 60
WBC<=20
WBC>20
39 41
Months 60
WBC<=20
WBC>20
Figure 1. Eect of white blood cell count on prognosis of AML (8;21). Probability of relapse in complete remittors with AML t(8;21) (panel A) and complete responders with AML without favourable cytogenetics (panel B). Overall survival for patients with AML t(8;21) is depicted in panel C, and for those without favourable cytogenetics in panel D. A white blood cell count of 20 109/l or more has a negative impact in the AML t(8;21) subgroup but not in other patients with AML. Thus the use of a WBC count criterion separates a relatively poor risk subset (20% of cases) from a cytogenetically good risk AML t(8;21). N, number of cases; O, number of events.
48 8
N O WBC<=20 63 21 WBC>20 14 9 Log rank P = 0.02
(C) t(8;21)
At risk: WBC<=20 63 WBC>20 14
0
25
50
75
100
41 6
N O WBC<=20 58 12 WBC>20 13 5 Log rank P = 0.05
At risk: WBC<=20 58 WBC>20 13
Relapse free (%)
Overall survival (%)
Relapse free (%) Overall survival (%)
(A) t(8;21)
68 B. LoÈwenberg
Prognostic factors in acute myeloid leukaemia 69
WHITE BLOOD CELL COUNT In several large prospective studies of previously untreated patients with AML, the WBC count stands out as an independent prognostic factor. There is a subgroup of patients presenting at diagnosis with hyperleucocytosis (WBC more than 100 109/l) who generally have a signi®cantly reduced complete response rate and a greater rate of relapse.22 The WBC count also provides a practically useful parameter for distinguishing the prognostic heterogeneity among patients with favourable cytogenetics. Thus those with t(8;21), inv(16) and t(15;17) as well as a WBC of 20 109/l or greater do signi®cantly worse than those with the same cytogenetic subtypes and low WBC counts (Figure 1). FAB CLASSIFICATION In multivariate analysis, the cytomorphological classi®cation also appears to add some prognostic information, independent of cytogenetics and molecular genotyping. In particular AMLs with the FAB types M0, M6 and M7 appear to correlate with an inferior outcome in most of the large phase III prospective trials. Since patients with M0, M6 and M7 represent minorities, the basis of the evidence for their prognostic value has remained limited and in clinical practice FAB classi®cation is not generally applied to risk assessment in AML. In small series of patients with minimally dierentiated AML (FAB M0), AML-M0 was associated with a poor prognosis.23±27 Also AML with the natural killer (NK) cell immunophenotype appears to carry a poor prognosis.28 Notably, a signi®cant proportion of AML-M0 appear to carry point mutations in the runt domain of the AML1 gene. The functional consequences of these mutations in critical myeloid transcription factors are currently under investigation.29 DRUG RESISTANCE Resistance of AML to the cytotoxic eects of chemotherapy has remained the main stumbling block in obtaining a cure. Thus one would assume that insights into the molecular pathways leading to drug resistance might provide particularly powerful prognostic markers for response to therapy. Multidrug resistance type I (MDR1) or classical drug resistance is associated with the enhanced expression of P-glycoprotein (Pgp), an ATP binding drug transporter in the plasma membrane of leukaemic cells. This transporter may bind a variety of substrates (including anthracyclins and epipodophylotoxins). It acts as an eux pump for these drugs. High levels of MDR1 have been associated with reduced intracellular concentrations of chemotherapy within leukaemic cells (see Chapter 12). The frequency of MDR1 positive AML markedly increases with age.14,30 In several studies MDR1 positivity has been shown to have a negative prognostic value for response to induction chemotherapy.14,30±35 This was seen in studies in which MDR1 was assessed by Pgp staining or by mRNA measurements. Other studies did not ®nd such correlations.36 It should be noted that a lack of agreement between some studies may relate to technical variations, e.g. the use of dierent endpoints for positivity, dierences in cell sample preparation (e.g. fresh versus cryopreserved) and analysis, and the use of dierent immunological reagents.37 In some studies functional assays of drug export (e.g. rhodamine export inhibitable by an MDR blocker) have
70 B. LoÈwenberg
been used and have shown prognostic signi®cance for response.30,38,39 Of particular interest are the results of a study indicating a strong prognostic impact of simultaneous functional measurements of multidrug resistance associated protein (MRPI) and Pgp activity with regard to response.38 Combined MRP1 and Pgp resistance was mainly seen among the poor cytogenetic subgroup of AML. Notably, in most of these studies the prognostic signi®cance of MDR1 has been evaluated in the context of daunomycin chemotherapy. One study of patients treated with induction chemotherapy using idarubicin did not demonstrate a correlation between MDR1 expression and response.40 It is not unlikely that multiple resistance mechanisms co-operate in the same patient and in synergy determine clinical resistance. This is supported by the recent observation of two distinct resistance phenotypes that in combination added to the prognostic impact of the individual marker.38 Finally, it would probably make more sense to relate resistance phenotyping to speci®c subpopulations of AML cells, i.e. AML progenitor cells, rather than the overall AML cell population. In one study the assessment of MDR1 expression on the subpopulation of CD34 cells by dual surface marker analysis, signi®cantly enhanced the predictive value for response.34 Other genes involved in the cellular redistribution of chemotherapeutic agents are MRP (MDR associated protein), a transporter of the glutathione complex and the lung resistance protein (LRP). Some studies have indicated that LRP may predict for response as well as for leukaemia-free survival39,42,43, but other studies did not reveal a correlation between the expression of LRP and response.30,43
PROLIFERATIVE ABILITIES IN VITRO AND GROWTH FACTOR RECEPTOR MUTATIONS It has been shown that autonomous proliferation of AML cells in short term culture predicts for a comparatively high probability of relapse and poor survival.44 Patients with AML and high spontaneous proliferative activity, also more frequently express CD34 positivity and a multidrug resistance phenotype.34 More recently mutations in the receptors for the haematopoietic growth factors have been detected, in particular in fms-like tyrosine kinase 3 (¯t-3, receptor for FL or ¯t ligand) and kit (receptor for stem cell factor or kit-ligand).22,45±51 Flt-3, kit and fms (receptor for M-CSR) are class III receptor tyrosine kinases, which play a role in haematopoiesis as growth factor receptors and share similar structural domains. In a signi®cant proportion of cases of AML, c-kit50,52,53 and c-fms54 are expressed. A subset of cases of AML carry mutations of c-kit and c-fms. Kit mutations are mainly seen in patients with good risk cytogenetics.55 Internal tandem duplicates (ITDs) of ¯t-3 have been reported in the part of the ¯t-3 gene coding for the juxtamembrane (JM) in 15±20% of AML patients.22,51 The elongation of the JM domain causes ligand independent dimerization thus resulting in constitutive activation and a reduced responsiveness to stromal support in long term culture.49 Flt-3 mutations have been shown to confer a grave prognosis both in young and older adults, independent of other prognostic factors. Of particular note is that the large subset of cytogenetically de®ned patients of intermediate risk can be split into good risk and poor risk according the absence or presence of ¯t-3 mutations51 (Figure 2).
Prognostic factors in acute myeloid leukaemia 71
1.0
(A)
0.8
0.6 n=95 0.4
0.2 Event-free survival
n=29 0.0
0
1.0
12
24
36
48
60
(B)
0.8
0.6 n=69
0.4
0.2
0.0
n=26 0
12
24
36
48
60
Time after diagnosis (months) Figure 2. Kaplan±Meier event-free survival curves of the eect of Flt-3 mutations in AML. Continuous lines indicates Flt-3/ITD (internal tandem duplication) negative samples. Dotted lines refer to Flt-3/ITD positive populations. (A) gives event-free survival of the total AML population. (B) depicts event-free survival of the AML population with intermediate-risk cytogenetics.
SECONDARY LEUKAEMIA AND ADVANCED DISEASE There is a considerable body of data indicating that patients with AML subsequent to an antecedent haematological disorder, e.g. myelodysplasia, polycythemia vera, congenital neutropenia have a reduced probability of attaining a complete response, and disease-free survival and overall survival are also reduced.14,56
72 B. LoÈwenberg Table 1. Major prognostic factors in acute myeloid leukemia. For response
For relapse
Cytogenetics/molecular genetics
Cytogenetics/molecular genetics Time towards complete response White blood cell count ¯t-3 mutations Autonomous proliferation Secondary AML Age
White blood cell count MDR phenotype Secondary AML Age
Patients with AML relapsing after an initial complete response, have a notoriously bad prognosis overall. By and large, less than 10% of these will have the prospect of long-term leukaemia-free survival. Usually these patients are entered on to transplant protocols or, if these options are not available, they may be oered experimental therapy.57 A small subset (5±10%) of patients with a distinctly better prognosis may be identi®ed among cases of recurrent AML. These are those who have favourable cytogenetics (t(8;21), inv 16, t(15;17)), relapse following a ®rst remission of at least 12 months duration, have not had a prior stem cell transplant and are less than 35 years old. In particular, patients with recurrent AML with at least three of the latter characteristics have been estimated to have a probability of 5 years disease-free survival of approximately 40%.
CONCLUSION The understanding of dominant prognostic determinants of response to therapy in patients with AML is rapidly evolving. Cytogenetics has become part of the essential and standard work-up of patients with AML and currently furnishes distinctive insights into the nature of the disease and provides a useful clue to the prognosis of individual patients. However more precise distinctions are needed. New parameters with powerful prognostic signi®cance are emerging. When these latter variables are considered in one comprehensive prognostic model and when their value has been validated in large series of patients, these parameters are likely to provide more exact quantitative estimations of the prognosis. These prognostic distinctions are likely to provide the elementary foundations for treatment choice in the near future.
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Internal tandem publication of the Flt-3 gene is preferentially seen in acute myeloid leukemia and myelodysplastic syndrome among various hematological malignancies: a study on a large series of patients and cell lines. Leukemia 1997; 11: 1605±1609. 47. Horiike S, Yokota S, Nakao M et al. Tandem duplications of the Flt-3 receptor gene are associated with leukemic transformation of myelodysplasia. Leukemia 1997; 11: 1442±1446. 48. Kiyoi H, Naoe T, Yokota S, Nakao M & Minami S. Internal tandem duplication of Flt-3 associated with leukocytosis in acute promyelocytic leukemia. Leukemia 1997; 11: 1447±1452. 49. Rombouts WC, Broyl A, Martens ACM, Slater R & Ploemacher R. Human acute myeloid leukemia cells with internal tandem duplications in the Flt-3 gene show reduced proliferative ability in stroma supported long-term cultures. Leukemia 1999; 13: 1071±1078. 50. Gari M, Goodeye A, Wilson G et al. c-Kit protooncogene exon 8 in-frame deletion plus insertion mutations in acute myeloid leukemia. British Journal of Haematology 1999; 105: 894±900. 51. Rombouts WJC, Blokland I, LoÈwenberg B & Ploemacher RE. Biological characteristics and prognosis of adult acute myeloid leukemia with internal tandem duplications in the Flt3 gene. Leukemia 2000; 14: 675±683.
Prognostic factors in acute myeloid leukaemia 75 52. Schwartz S, Heinecke A, Zimmermann M et al. Expression of the c-kit receptor (CD117) is a feature of almost all subtypes of de novo acute myeloblastic leukemia (AML), including cytogenetically good-risk AML, and lacks prognostic signi®cance. Leukemia and Lymphoma 1999; 34: 85±94. 53. Wang, Curtis JE, Geiseller EN, McCullock EA & Minden MD. The expression of the proto oncogene ckit in the blast cells of acute myeloblastic leukemia. Leukemia 1989; 3: 699±702. 54. Dubreuil P, Torres H, Courcoul MA, Birg F & Mannoni P. C-fms expression is a marker of human acute myeloid leukemias. Blood 1988; 72: 1081±1085. 55. Berghini A, Peterlongo P, Ripamonti CB, Larizza L & Cairoli R. C-kit mutations in core binding leukemias. Blood 2000; 95: 726±727. 56. Keating AJ, McCredie KB, Benjamin RS et al. Treatment of patients over 50 years of age with acute myelogenous leukemia with a combination of rubidizone and cytosine, arabinoside, vincristine and prednisone (ROAP). Blood 1981; 58: 584±590. 57. LoÈwenberg B, Downing JR & Burnett A. Acute myeloid leukemia. New England Journal of Medicine 1999; 341: 1051±1962. 58. Osato M, Asou N, Abdalla E et al. Bi-allelic and heterozygous point mutation in the runt domain of the AML1/PEBP2aB gene associated with acute myeloblastic leukemia. Blood 1999; 93: 1817±1824. 59. Cahn JY, Labopin M, Mandelli F et al. Autologous bone marrow transplantation for ®rst remission acute myeloblastic leukemia in patients older than 50 years; a retrospective analysis of the European Bone Marrow Transplant Group. Blood 1995; 85: 575±579.
doi:10.1053/beha.2000.0116, available online at http://www.idealibrary.com on
4 Prognostic factors in acute myeloid leukaemia Bob LoÈwenberg
MD, PhD
Professor of Haematology Erasmus University and University Hospital Rotterdam, P.O. Box 2040, 3000 CA Rotterdam, The Netherlands
The prognosis between subgroups of patients with acute myeloid leukaemia (AML) may range considerably. Haematological, genetic and clinical analysis has provided possibilities for de®ning the heterogeneity of prognosis. This furnishes clinically relevant insights into the probability of treatment response and survival in individual cases of AML and it provides a lead in treatment management. Key words: prognosis; prognostic factors; acute myeloid leukaemia; cytogenetics; ¯t-3 receptor; ¯t-3 mutations; molecular genetics.
INTRODUCTION The group of diseases collectively designated as acute myeloid leukaemia (AML) represents a broad variety of distinct entities. AML arises from a malignantly transformed haematopoietic stem cell or progenitor cell. The cell stage (more or less primitive, multipotent or committed) at which transformation occurs and the residual capabilities of maturation of these cells, may contribute to the phenotypic diversity of this disease.1±3 These variations may be re¯ected in the dierent cytomorphology of the leukaemic blasts, the dysplasia of the other haematopoietic cell lineages and the characteristic sites of presentation of leukaemia (e.g. skin, gingiva etc). The molecular abnormalities leading to full leukaemic transformation vary greatly. Dierences in the molecular pathogenesis may also dictate variation in clinical presentation as well as variation of response to therapy. The understanding that the term AML lumps together a variety of distinct pathogenic conditions, has led to a general appreciation of the importance of delineating AML disease entities more sharply, e.g. AML with t(8;21) or AML1/eight-twenty-one (ETO) fusion gene etc. Each of the genetically de®ned AML diseases carries a typical prognosis and one may anticipate that in the long run these conditions will require speci®c therapy. Acute promyelocytic leukaemia with t(15;17) and the promyelocytic leukaemia gene/retinoic acid receptor alpha gene (PML/RAR-a) fusion gene, treated with all-trans retinoic acid (ATRA) is a classic example of this approach.4 Meanwhile a thorough understanding of the individual prognostic factors has become of signi®cant importance for treatment choice in today's clinical practice. Should the patient be treated with an allograft from an HLA-identical sibling or should 1521±6926/01/01006511 $35.00/00
c 2001 Harcourt Publishers Ltd. *
66 B. LoÈwenberg
the transplant be withheld and reserved in case of relapse? Who should receive an allogeneic HLA matched transplant from an unrelated donar and when? Who should receive high-dose cytarabine as part of their chemotherapy? Who should be entered onto experimental protocols? In addition to cytogenetics in recent years new leukaemia-related prognostic factors (e.g. ¯t-3 mutations) have been recognized, which may have an important impact on prognosis. Here we will review the current knowledge of factors that are predictive of the response to therapy and outcome in adults with AML (Table 1). AGE AND GENDER Age has a strong prognostic impact on the outcome of AML. There is a gradual but consistent decline in the prognosis for patients with AML with increasing age. This negative relationship is continuous from infant to 85 years of age. The latter relationship is apparent as a progressive decrease in treatment response and survival as a function of decade of age (Oxford Collaborative AML Intergroup, pers. comm.). For practical reasons of protocol design, a cut-o point of 60 years is usually used to distinguish older patients. Young and middle-aged adults, i.e. those less than 60 years old, have response rates to remission induction chemotherapy of approximately 75% and approximately 35±40% of them survive following diagnosis for 5 years or more.5±11 In contrast, patients with AML who are more than 60 years old have an overall response probability to induction therapy of 45±55%, and fewer than 10% of these survive for a minimum of 5 years.12,13 While the results of treatment have improved steadily over the last 20 years in younger adults, due to more intensive cytotoxic treatment, the use of stem cell grafts and improvements in supportive care, no signi®cant change in outcome has been noted among individuals of 60 years of age. The outcome for older subjects may even be worse than these estimates would indicate, since many older patients are not referred to hospitals for treatment, and are not registered or evaluated for outcome. Thus these outcome data for aged individuals probably represent overestimations. Why is outcome so unsatisfactory in the aged? The reasons for the poorer outcome of patients of higher age most probably relate to the increased frequency of unfavourable cytogenetics among older patients with AML, a greater frequency of antecedent myelodysplasia, as well as a greater frequency of drug resistance phenotypes. Also, because of the reduced general health of older patients, they cannot withstand intensive chemotherapy as well as younger individuals do.13,14 Several large databases of phase III studies in untreated patients with AML would also suggest that female sex is an independent favourable prognostic parameter for disease-free survival, although of moderate impact only. The intergroup analysis of the Oxford Collaborative AML Intergroup (pers. comm.) has con®rmed that disease-free survival of male patients is somewhat less than that of females. CYTOGENETICS AND MOLECULAR GENETICS Cytogenetic abnormalities are seen in approximately 60% of cases in AML and are highly predictive of response to therapy as well as the probability of relapse.15±18 The translocations t(8;21), inv16 and t(15;17) generally carry a relatively favourable prognosis and these are more commonly seen among younger patients with AML. The fusion genes of each of these translocations have been identi®ed and can be detected
Prognostic factors in acute myeloid leukaemia 67
using molecular probes in reverse transcriptase-polymerase chain reactions (RT-PCR). Among patients with these chromosomal abnormalities or the corresponding molecular abnormalities (AML1-ETO, CBFb-MYH11 (core binding factor beta genesmooth muscle myosin heavy chain gene), PML-RARa) the response to induction therapy is 80% or greater and for those entering remission the probability of relapse is 30±40% resulting in 5 years survival rates of 60±70%. The latter molecular or cytogenetic subsets of AML correlate with certain cytomorphological categories. For instance, t(15;17) or PML/RAR correlates with the French±American±British (FAB) group subtype M3, and inv16 with cytological FAB subtype M4 with eosinophils. However, the cytogenetics and molecular genetics rather than the cytological FAB classes, determine the overriding prognostic signi®cance. In contrast patients with t(6;9), 11q23 (MLL-gene) abnormalities19, monosomies of chromosomes 7 (-7) or 5 (-7) and complex cytogenetic abnormalities (three dierent cytogenetic aberrations or more) generally have a distinctively poor prognosis. In these individuals (when 60 years or younger), the average complete response to induce treatment is 60% with a majority (approximately 80%) relapsing within 2 years so that their survival at 5 years is approximately 15% only.11,20 The cytogenetic and molecular indicators have led to the de®nition of prognostic groups, i.e. favourable risk (with favourable molecular genetics or cytogenetics), unfavourable risk (those with poor-risk cytogenetics) and intermediate risk (all others). Usually, these risk classi®cations are re®ned by considering selected additional prognostic determinants that enhance the value of these cytogenetic or molecular distinctions. The risk-scoring systems that are based upon multiple prognostic factors may permit a more precise approximation of prognosis in individual cases. To keep this practically simple, most collaborative groups have restricted the use of these additional parameters, and only taken into account a limited number of covariables that have a considerable impact. As an example, the Dutch HOVON (Haemato-Oncology Collaborative Group for Adults±Netherlands) group has required a white blood cell (WBC) count of less than 20 109/l in addition to favourable t(8;21) cytogenetics for the good risk category assignment (risk of relapse is 24% at 5 years: Figure 1). The Medical Research Council (MRC) has included the rapid versus late attainment of complete response (i.e. after cycle 1 versus after cycle 2) in their risk score in order to enhance the separation between good risk and poor risk.20
IMMUNOPHENOTYPING Cell stage and cell lineage speci®c markers have been used for immunophenotyping of AML. While the cytogenetic and molecular genetic markers carry a strong prognostic impact, the value of immunophenotyping for the assessment of prognosis has not been ®rmly established. This is true for the myeloid markers CD13, CD14, CD15, CD11b, HLA-DR, CD117 (c-kit) as well as CD34 for which controversial results regarding the prognostic value for response have been obtained in a variety of studies.21 The choice of antibodies, the threshold values for positivity (e.g. percentage positive cells) and clinical variations may explain some of the discrepancy between dierent studies. More importantly, most positive correlations between immunophenotype and prognosis do not hold up in multivariate analysis indicating that other associated determinants exert a greater eect on prognosis.
0
25
50
75
100
0
0 39 4
34 4
27 3
20 2
23 2
Months 60
WBC>20
WBC<=20
24 3
Months 60
WBC>20
WBC<=20
0 96 79
0 297 206
N O WBC<=20 686 422 WBC>20 570 344 Log rank P = 0.27
152 100
(D) No favourable cytogenetics
At risk: WBC<=20 686 WBC>20 570
0
25
50
75
100
182 147
N O WBC<=20 546 269 WBC>20 450 203 Log rank P = 0.82
At risk: WBC<=20 546 WBC>20 450
0
25
50
75
100
(B) No favourable ctyogenetics
91 71
65 58
56 49
Months 60
WBC<=20
WBC>20
39 41
Months 60
WBC<=20
WBC>20
Figure 1. Eect of white blood cell count on prognosis of AML (8;21). Probability of relapse in complete remittors with AML t(8;21) (panel A) and complete responders with AML without favourable cytogenetics (panel B). Overall survival for patients with AML t(8;21) is depicted in panel C, and for those without favourable cytogenetics in panel D. A white blood cell count of 20 109/l or more has a negative impact in the AML t(8;21) subgroup but not in other patients with AML. Thus the use of a WBC count criterion separates a relatively poor risk subset (20% of cases) from a cytogenetically good risk AML t(8;21). N, number of cases; O, number of events.
48 8
N O WBC<=20 63 21 WBC>20 14 9 Log rank P = 0.02
(C) t(8;21)
At risk: WBC<=20 63 WBC>20 14
0
25
50
75
100
41 6
N O WBC<=20 58 12 WBC>20 13 5 Log rank P = 0.05
At risk: WBC<=20 58 WBC>20 13
Relapse free (%)
Overall survival (%)
Relapse free (%) Overall survival (%)
(A) t(8;21)
68 B. LoÈwenberg
Prognostic factors in acute myeloid leukaemia 69
WHITE BLOOD CELL COUNT In several large prospective studies of previously untreated patients with AML, the WBC count stands out as an independent prognostic factor. There is a subgroup of patients presenting at diagnosis with hyperleucocytosis (WBC more than 100 109/l) who generally have a signi®cantly reduced complete response rate and a greater rate of relapse.22 The WBC count also provides a practically useful parameter for distinguishing the prognostic heterogeneity among patients with favourable cytogenetics. Thus those with t(8;21), inv(16) and t(15;17) as well as a WBC of 20 109/l or greater do signi®cantly worse than those with the same cytogenetic subtypes and low WBC counts (Figure 1). FAB CLASSIFICATION In multivariate analysis, the cytomorphological classi®cation also appears to add some prognostic information, independent of cytogenetics and molecular genotyping. In particular AMLs with the FAB types M0, M6 and M7 appear to correlate with an inferior outcome in most of the large phase III prospective trials. Since patients with M0, M6 and M7 represent minorities, the basis of the evidence for their prognostic value has remained limited and in clinical practice FAB classi®cation is not generally applied to risk assessment in AML. In small series of patients with minimally dierentiated AML (FAB M0), AML-M0 was associated with a poor prognosis.23±27 Also AML with the natural killer (NK) cell immunophenotype appears to carry a poor prognosis.28 Notably, a signi®cant proportion of AML-M0 appear to carry point mutations in the runt domain of the AML1 gene. The functional consequences of these mutations in critical myeloid transcription factors are currently under investigation.29 DRUG RESISTANCE Resistance of AML to the cytotoxic eects of chemotherapy has remained the main stumbling block in obtaining a cure. Thus one would assume that insights into the molecular pathways leading to drug resistance might provide particularly powerful prognostic markers for response to therapy. Multidrug resistance type I (MDR1) or classical drug resistance is associated with the enhanced expression of P-glycoprotein (Pgp), an ATP binding drug transporter in the plasma membrane of leukaemic cells. This transporter may bind a variety of substrates (including anthracyclins and epipodophylotoxins). It acts as an eux pump for these drugs. High levels of MDR1 have been associated with reduced intracellular concentrations of chemotherapy within leukaemic cells (see Chapter 12). The frequency of MDR1 positive AML markedly increases with age.14,30 In several studies MDR1 positivity has been shown to have a negative prognostic value for response to induction chemotherapy.14,30±35 This was seen in studies in which MDR1 was assessed by Pgp staining or by mRNA measurements. Other studies did not ®nd such correlations.36 It should be noted that a lack of agreement between some studies may relate to technical variations, e.g. the use of dierent endpoints for positivity, dierences in cell sample preparation (e.g. fresh versus cryopreserved) and analysis, and the use of dierent immunological reagents.37 In some studies functional assays of drug export (e.g. rhodamine export inhibitable by an MDR blocker) have
70 B. LoÈwenberg
been used and have shown prognostic signi®cance for response.30,38,39 Of particular interest are the results of a study indicating a strong prognostic impact of simultaneous functional measurements of multidrug resistance associated protein (MRPI) and Pgp activity with regard to response.38 Combined MRP1 and Pgp resistance was mainly seen among the poor cytogenetic subgroup of AML. Notably, in most of these studies the prognostic signi®cance of MDR1 has been evaluated in the context of daunomycin chemotherapy. One study of patients treated with induction chemotherapy using idarubicin did not demonstrate a correlation between MDR1 expression and response.40 It is not unlikely that multiple resistance mechanisms co-operate in the same patient and in synergy determine clinical resistance. This is supported by the recent observation of two distinct resistance phenotypes that in combination added to the prognostic impact of the individual marker.38 Finally, it would probably make more sense to relate resistance phenotyping to speci®c subpopulations of AML cells, i.e. AML progenitor cells, rather than the overall AML cell population. In one study the assessment of MDR1 expression on the subpopulation of CD34 cells by dual surface marker analysis, signi®cantly enhanced the predictive value for response.34 Other genes involved in the cellular redistribution of chemotherapeutic agents are MRP (MDR associated protein), a transporter of the glutathione complex and the lung resistance protein (LRP). Some studies have indicated that LRP may predict for response as well as for leukaemia-free survival39,42,43, but other studies did not reveal a correlation between the expression of LRP and response.30,43
PROLIFERATIVE ABILITIES IN VITRO AND GROWTH FACTOR RECEPTOR MUTATIONS It has been shown that autonomous proliferation of AML cells in short term culture predicts for a comparatively high probability of relapse and poor survival.44 Patients with AML and high spontaneous proliferative activity, also more frequently express CD34 positivity and a multidrug resistance phenotype.34 More recently mutations in the receptors for the haematopoietic growth factors have been detected, in particular in fms-like tyrosine kinase 3 (¯t-3, receptor for FL or ¯t ligand) and kit (receptor for stem cell factor or kit-ligand).22,45±51 Flt-3, kit and fms (receptor for M-CSR) are class III receptor tyrosine kinases, which play a role in haematopoiesis as growth factor receptors and share similar structural domains. In a signi®cant proportion of cases of AML, c-kit50,52,53 and c-fms54 are expressed. A subset of cases of AML carry mutations of c-kit and c-fms. Kit mutations are mainly seen in patients with good risk cytogenetics.55 Internal tandem duplicates (ITDs) of ¯t-3 have been reported in the part of the ¯t-3 gene coding for the juxtamembrane (JM) in 15±20% of AML patients.22,51 The elongation of the JM domain causes ligand independent dimerization thus resulting in constitutive activation and a reduced responsiveness to stromal support in long term culture.49 Flt-3 mutations have been shown to confer a grave prognosis both in young and older adults, independent of other prognostic factors. Of particular note is that the large subset of cytogenetically de®ned patients of intermediate risk can be split into good risk and poor risk according the absence or presence of ¯t-3 mutations51 (Figure 2).
Prognostic factors in acute myeloid leukaemia 71
1.0
(A)
0.8
0.6 n=95 0.4
0.2 Event-free survival
n=29 0.0
0
1.0
12
24
36
48
60
(B)
0.8
0.6 n=69
0.4
0.2
0.0
n=26 0
12
24
36
48
60
Time after diagnosis (months) Figure 2. Kaplan±Meier event-free survival curves of the eect of Flt-3 mutations in AML. Continuous lines indicates Flt-3/ITD (internal tandem duplication) negative samples. Dotted lines refer to Flt-3/ITD positive populations. (A) gives event-free survival of the total AML population. (B) depicts event-free survival of the AML population with intermediate-risk cytogenetics.
SECONDARY LEUKAEMIA AND ADVANCED DISEASE There is a considerable body of data indicating that patients with AML subsequent to an antecedent haematological disorder, e.g. myelodysplasia, polycythemia vera, congenital neutropenia have a reduced probability of attaining a complete response, and disease-free survival and overall survival are also reduced.14,56
72 B. LoÈwenberg Table 1. Major prognostic factors in acute myeloid leukemia. For response
For relapse
Cytogenetics/molecular genetics
Cytogenetics/molecular genetics Time towards complete response White blood cell count ¯t-3 mutations Autonomous proliferation Secondary AML Age
White blood cell count MDR phenotype Secondary AML Age
Patients with AML relapsing after an initial complete response, have a notoriously bad prognosis overall. By and large, less than 10% of these will have the prospect of long-term leukaemia-free survival. Usually these patients are entered on to transplant protocols or, if these options are not available, they may be oered experimental therapy.57 A small subset (5±10%) of patients with a distinctly better prognosis may be identi®ed among cases of recurrent AML. These are those who have favourable cytogenetics (t(8;21), inv 16, t(15;17)), relapse following a ®rst remission of at least 12 months duration, have not had a prior stem cell transplant and are less than 35 years old. In particular, patients with recurrent AML with at least three of the latter characteristics have been estimated to have a probability of 5 years disease-free survival of approximately 40%.
CONCLUSION The understanding of dominant prognostic determinants of response to therapy in patients with AML is rapidly evolving. Cytogenetics has become part of the essential and standard work-up of patients with AML and currently furnishes distinctive insights into the nature of the disease and provides a useful clue to the prognosis of individual patients. However more precise distinctions are needed. New parameters with powerful prognostic signi®cance are emerging. When these latter variables are considered in one comprehensive prognostic model and when their value has been validated in large series of patients, these parameters are likely to provide more exact quantitative estimations of the prognosis. These prognostic distinctions are likely to provide the elementary foundations for treatment choice in the near future.
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