Leukemic cells from a significant number of children with acute lymphoblastic leukemia (ALL) express protein antigens characteristic of both lymphoid and myeloid cells, yet the clinical significance of this immunophenotype has remained controversial. In the current study, we have determined relationships between myeloid antigen expression and treatment outcome in a large cohort of children with newly diagnosed ALL. A total of 1,557 children enrolled on risk-adjusted Children's Cancer Group studies were classified as myeloid antigen positive (My+) or myeloid antigen negative (My) B-lineage ALL (BL) or T-lineage ALL (TL), according to expression of CD7, CD19, CD13, and CD33 antigens on the surface of their leukemic cells. My+ patients in both BL and TL groups were more likely than My patients to have favorable presenting features. Induction therapy outcome was similar for My+ and My patients in both the BL and TL categories. Importantly, 4-year event-free survival (EFS) was similar for My+ BL (77.0%, standard deviation [SD] = 4.0%) versus My BL (75.9%, SD = 1.8%) and for My+ TL (72.7%, SD = 7.1%) versus My TL (70.1%, SD = 5.7%). An overall relative hazard rate (RHR) of 0.89 (P = .49) was determined by a cross strata analysis for My+ versus My patients. Moreover, similar EFS and RHR also were found when My+ and My BL patients were compared according to National Cancer Institute risk classification. Thus, patients with My+ ALL have similar treatment outcomes as My ALL patients. In contrast to previous studies, this result was independent of treatment risk category, demonstrating that myeloid antigen expression was not an adverse prognostic factor for childhood ALL.

ACUTE LYMPHOBLASTIC leukemia (ALL) is an immunophenotypically heterogeneous group of diseases. Leukemic cells from the majority of patients with ALL express on their surface a variety of protein antigens that are found at discrete stages of maturation on normal B- or T-lymphocyte precursors.1-6 Thus leukemic clones from ALL patients are thought to originate from normal lymphoid progenitor cells arrested at early stages of B- or T-lymphocyte ontogeny.7-9 Recent improvements in immunofluorescence and flow cytometry, as well as the availability of monoclonal antibodies that recognize lineage-associated cell surface molecules, have motivated more detailed investigations of immunophenotypic heterogeneity in childhood ALL. It is now clear that leukemic cells from a 5% to 20% of children with ALL also express myeloid differentiation antigens.5,10-20 The expression of myeloid antigens by ALL cells is speculated to reflect either lineage infidelity due to aberrant gene expression, neoplastic transformation of rare bilineage lymphoid/myeloid progenitor cells, or transformation of a multipotent lymphohematopoietic precursor cell.6 21-23 

The clinical significance of myeloid antigen expression in pediatric ALL has remained controversial. Several studies5,14,15,20 have reported poor outcome for children with ALL of mixed myeloid/lymphoid phenotype, whereas others have found similar induction and treatment outcomes for patients with myeloid antigen negative and myeloid antigen positive ALL.10,11,13,16-19 24 Because of these conflicting reports regarding prognosis and treatment outcome, there has been no consensus among pediatric oncologists regarding assignment of patients to risk-directed ALL chemotherapy protocols, employment of therapies directed at acute myelogenous leukemia (AML), or necessity for bone marrow transplantation in first remission.

Herein, we report the results of a prospective study of myeloid antigen expression in a large cohort of 1,557 children with newly diagnosed ALL who were enrolled on risk-adjusted treatment protocols of the Children's Cancer Group (CCG). The presenting features and treatment outcomes of both myeloid antigen positive (My+) B-lineage (BL) and My+ T-lineage (TL) ALL patients were compared with those of myeloid antigen negative (My) BL and My TL controls. Our results provide new insights regarding the clinical relevance of myeloid antigen expression in childhood ALL by demonstrating that regardless of treatment protocol, My+ ALL and My ALL patients have similar treatment outcomes.

Study patients.The sample for these analyses included pediatric patients (<21 years of age) with newly diagnosed ALL enrolled between January 1, 1989 and December 31, 1993 on risk-adjusted treatment protocols of the CCG for whom a complete immunophenotyping profile of specified lymphoid and myeloid antigens (see below) was obtained. Diagnosis of ALL was based on morphological, biochemical, and immunological features of the leukemic cells, including lymphoblast morphology on Wright-Giemsa stained bone marrow smears, positive nuclear staining for terminal deoxynucleotidyl transferase (TdT), negative staining for myeloperoxidase, and cell surface expression of two or more lymphoid differentiation antigens (see below). Degree of organomegaly (moderate or marked enlargement) was as defined previously.25 CCG risk-adjusted ALL protocols were as follows: CCG 1881 (low-risk protocol for children age 2 to 9 years and white blood cell count [WBC] < 10,000/μL); CCG 1882 (high-risk protocol for patients 1 to 9 years of age with WBC ≥ 50,000/μL or age ≥ 10 years); CCG 1883 (protocol for infants less than 1 year of age), 1891 (intermediate risk protocol for children aged 2 to 9 years and WBC 10,000 to 49,999/μL or age 1 year and WBC < 50,000/μL) and CCG 1901 (high-risk protocol for patients with lymphomatous features). Lymphomatous features are essentially as described by the revised criteria of Steinherz et al.25 Each protocol was approved by the National Cancer Institute (NCI), as well as the Institutional Review Boards of the participating CCG-affiliated institutions. Informed consent was obtained from parents, patients, or both, as deemed appropriate, according to Department of Health and Human Services guidelines. For comparisons of presenting features, antigen expression, and therapy outcomes, patients were classified as myeloid antigen positive (My+) B-lineage leukemia (BL), myeloid antigen negative (My) BL, My+ T-lineage leukemia (TL) and My TL, as described below. A small number of patients (24 BL and 3 TL) were excluded from the current analyses because they failed to meet the criteria given by the algorithm. Analyses performed using these 27 patients indicated similar presenting characteristics and outcome compared with the patients included in this report. Thus, there appears to be no selection bias associated with the removal of these patients. B-lineage My+ and My patients were also grouped according to recently published NCI risk classification criteria.26 These criteria classify patients age 1 to 9 years and WBC < 50,000/μL as standard risk and all other patients as high-risk.

Table 1.

Antigen Expression in Children With My+ and My ALL

SurfaceBL
AntigenMy+ BLMy BLP ValueSurface AntigenTL
No.Mean (±SE)*Med.No.Mean (±SE)Med.My+ TLMy TLP Value
No.Mean (±SE)Med.No.Mean (±SE)Med.
CD19 217 89.0 (0.5) 91 1,113 86.6 (0.4) 91 .57 CD2 42 68.6 (5.6) 90 184 76.7 (2.2) 92 .61 
CD13 217 49.1 (1.6) 47 1,113 4.7 (0.2) NA CD5 43 58.7 (6.1) 78 184 85.9 (1.4) 93 .001 
CD33 217 28.8 (2.0) 19 1,113 2.4 (2.3) NA CD7 43 89.3 (1.7) 94 184 89.0 (0.9) 94 .90 
CD10 217 77.2 (1.8) 88 1,113 77.2 (0.8) 88 .86 CD10 43 21.9 (5.1) 184 26.5 (2.6) .25 
CD34 214 68.8 (1.9) 78 1,106 53.7 (1.1) 64 <.0001 CD13 43 52.2 (4.6) 51 184 3.7 (0.4) NA 
CD40 190 68.6 (2.1) 80 860 48.8 (1.1) 53 <.0001 CD33 43 21.0 (4.1) 184 2.2 (0.3) NA 
        CD34 43 52.5 (5.1) 60 183 30.0 (2.6) .0002 
SurfaceBL
AntigenMy+ BLMy BLP ValueSurface AntigenTL
No.Mean (±SE)*Med.No.Mean (±SE)Med.My+ TLMy TLP Value
No.Mean (±SE)Med.No.Mean (±SE)Med.
CD19 217 89.0 (0.5) 91 1,113 86.6 (0.4) 91 .57 CD2 42 68.6 (5.6) 90 184 76.7 (2.2) 92 .61 
CD13 217 49.1 (1.6) 47 1,113 4.7 (0.2) NA CD5 43 58.7 (6.1) 78 184 85.9 (1.4) 93 .001 
CD33 217 28.8 (2.0) 19 1,113 2.4 (2.3) NA CD7 43 89.3 (1.7) 94 184 89.0 (0.9) 94 .90 
CD10 217 77.2 (1.8) 88 1,113 77.2 (0.8) 88 .86 CD10 43 21.9 (5.1) 184 26.5 (2.6) .25 
CD34 214 68.8 (1.9) 78 1,106 53.7 (1.1) 64 <.0001 CD13 43 52.2 (4.6) 51 184 3.7 (0.4) NA 
CD40 190 68.6 (2.1) 80 860 48.8 (1.1) 53 <.0001 CD33 43 21.0 (4.1) 184 2.2 (0.3) NA 
        CD34 43 52.5 (5.1) 60 183 30.0 (2.6) .0002 

Abbreviation: NA, not applicable.

*

Values are mean ± SE and median percentages of cells expressing the indicated antigen.

Kruskal-Wallis nonparametric rank test comparing My+ BL and My BL.

Kruskal-Wallis nonparametric rank test comparing My+ TL and My TL.

Immunophenotyping.Highly blast-enriched mononuclear cell fractions containing ≥90% leukemic cells were isolated from pretreatment bone marrow aspirate samples by centrifugation on Ficoll-Hypaque density gradients. Immunophenotyping was performed centrally in the CCG ALL Biology Reference Laboratory by indirect immunofluorescence and flow cytometry using monoclonal antibodies reactive with B-lymphoid–associated (CD19, CD20, CD21, CD22, CD72), T-lymphoid–associated (CD1, CD2, CD3, CD4, CD5, CD7, CD8), myeloid-associated (CD13, CD33), and nonlineage-associated (CD9, CD10, CD24, CD34, CD40) differentiation antigens, as previously described.27 28 Antigen expression data are presented as the mean ± standard error (SE) and median percentages of leukemic cells scored positive for expression of a given antigen. Cells were scored positive based on increased immunofluorescence observed with an antigen-specific monoclonal antibody compared with that observed with an irrelevant antibody. The term “expression frequency” is used throughout to indicate the percentage of leukemic cells expressing a given antigen. Patients were classified as BL if ≥30% of the isolated leukemic cells were positive for CD19 and < 30% were positive for CD2, CD5, and CD7. Likewise, patients were classified as TL if ≥30% of the isolated blasts were positive for CD2, CD5, or CD7 and <30% were positive for CD19. For patients exceeding 30% positivity for both criteria, the immunological surface marker results were examined further and classified according to the lineage marker of higher expression frequency, as well as the composite immunophenotype (ie, expression frequencies of other lineage-restricted antigens). BL and TL patients were classified as My+ if ≥30% of the isolated leukemic cells were positive for CD13 or CD33, or both. The majority of the 1,557 patients (85.4%) had BL, whereas 14.6% had TL. Overall, 13.9% patients were classified as My+ BL, 71.5% were My BL, 2.8% were My+ TL, and 11.8% were My TL.

Table 2.

Presenting Features of Children With BL According to Myeloid Antigen Expression

VariableCategoryMy+ BL (N = 217)My BL (N = 1,113)P Value*
No.(%)No.(%)
 
Age (yr) <1 (1.4) 48 (4.3) .12 
 1-9 169 (77.9) 845 (75.9)  
 ≥10 45 (20.7) 220 (19.8)  
WBC (×109/L) 1-19 149 (68.7) 647 (58.1) .006 
 20-49 24 (11.1) 208 (18.7)  
 ≥50 44 (20.3) 258 (23.2)  
Sex Male 123 (56.7) 616 (55.3) .72 
 Female 94 (43.3) 497 (44.7)  
Race White 166 (76.5) 836 (75.1) .83 
 Black 13 (6.0) 79 (7.1)  
 Other 38 (17.5) 198 (17.8)  
Down syndrome Yes (1.4) 25 (2.2) .58 
 No 214 (98.6) 1,087 (97.8)  
Liver Normal 115 (53.0) 504 (45.3) .12 
 Mod. enlarged 94 (43.3) 562 (50.5)  
 Markedly enlarged (3.7) 46 (4.1)  
Spleen Normal 123 (56.7) 476 (42.8) .0002 
 Mod. enlarged 88 (40.6) 551 (49.5)  
 Markedly enlarged (2.8) 86 (7.7)  
Lymph nodes Normal 138 (63.6) 572 (51.4) .004 
 Mod. enlarged 72 (33.2) 490 (44.0)  
 Markedly enlarged (3.2) 51 (4.6)  
Mediastinal mass Absent 215 (99.1) 1,076 (96.8) .16 
 Small (0.9) 30 (2.7)  
 Large (0.0) (0.5)  
Hemoglobin (g/dL) 1-7.9 133 (61.3) 656 (59.3) .86 
 8.0.-10.9 65 (30.0) 346 (31.3)  
 ≥11.0 19 (8.8) 104 (9.4)  
Platelets (×109/L) 1-49 96 (44.2) 590 (53.0) .01 
 50-149 66 (30.4) 331 (29.7)  
 ≥150 55 (25.4) 192 (17.3)  
CNS disease at diagnosis Yes (1.4) 27 (2.4) .35 
 No 212 (98.6) 1,081 (97.6)  
NCI risk category Standard 138 (64.2) 657 (59.9) .27 
 Poor 79 (35.8) 456 (40.1)  
Karyotypic features 
Number Diploid (46) 31 (43.7) 93 (24.1) .003 
 Hypodiploid (<46) (4.2) 30 (7.8)  
 Pseudodiploid (46) 20 (28.2) 95 (24.6)  
 Hyperdiploid (47-50) (11.3) 56 (14.5)  
 Hyperdiploid (>50) (12.7) 112 (29.0)  
Aberrations Normal 31 (43.7) 93 (24.1) .001 
 Abnormal 40 (53.6) 293 (75.9)  
Translocations t(4; 11) present (0.0) 13 (3.4) .12 
 t(4; 11) absent 71 (100.0) 373 (96.6)  
 t(9; 22) present (4.2) 10 (2.6) .45 
 t(9; 22) absent 68 (95.8) 376 (97.4) 
VariableCategoryMy+ BL (N = 217)My BL (N = 1,113)P Value*
No.(%)No.(%)
 
Age (yr) <1 (1.4) 48 (4.3) .12 
 1-9 169 (77.9) 845 (75.9)  
 ≥10 45 (20.7) 220 (19.8)  
WBC (×109/L) 1-19 149 (68.7) 647 (58.1) .006 
 20-49 24 (11.1) 208 (18.7)  
 ≥50 44 (20.3) 258 (23.2)  
Sex Male 123 (56.7) 616 (55.3) .72 
 Female 94 (43.3) 497 (44.7)  
Race White 166 (76.5) 836 (75.1) .83 
 Black 13 (6.0) 79 (7.1)  
 Other 38 (17.5) 198 (17.8)  
Down syndrome Yes (1.4) 25 (2.2) .58 
 No 214 (98.6) 1,087 (97.8)  
Liver Normal 115 (53.0) 504 (45.3) .12 
 Mod. enlarged 94 (43.3) 562 (50.5)  
 Markedly enlarged (3.7) 46 (4.1)  
Spleen Normal 123 (56.7) 476 (42.8) .0002 
 Mod. enlarged 88 (40.6) 551 (49.5)  
 Markedly enlarged (2.8) 86 (7.7)  
Lymph nodes Normal 138 (63.6) 572 (51.4) .004 
 Mod. enlarged 72 (33.2) 490 (44.0)  
 Markedly enlarged (3.2) 51 (4.6)  
Mediastinal mass Absent 215 (99.1) 1,076 (96.8) .16 
 Small (0.9) 30 (2.7)  
 Large (0.0) (0.5)  
Hemoglobin (g/dL) 1-7.9 133 (61.3) 656 (59.3) .86 
 8.0.-10.9 65 (30.0) 346 (31.3)  
 ≥11.0 19 (8.8) 104 (9.4)  
Platelets (×109/L) 1-49 96 (44.2) 590 (53.0) .01 
 50-149 66 (30.4) 331 (29.7)  
 ≥150 55 (25.4) 192 (17.3)  
CNS disease at diagnosis Yes (1.4) 27 (2.4) .35 
 No 212 (98.6) 1,081 (97.6)  
NCI risk category Standard 138 (64.2) 657 (59.9) .27 
 Poor 79 (35.8) 456 (40.1)  
Karyotypic features 
Number Diploid (46) 31 (43.7) 93 (24.1) .003 
 Hypodiploid (<46) (4.2) 30 (7.8)  
 Pseudodiploid (46) 20 (28.2) 95 (24.6)  
 Hyperdiploid (47-50) (11.3) 56 (14.5)  
 Hyperdiploid (>50) (12.7) 112 (29.0)  
Aberrations Normal 31 (43.7) 93 (24.1) .001 
 Abnormal 40 (53.6) 293 (75.9)  
Translocations t(4; 11) present (0.0) 13 (3.4) .12 
 t(4; 11) absent 71 (100.0) 373 (96.6)  
 t(9; 22) present (4.2) 10 (2.6) .45 
 t(9; 22) absent 68 (95.8) 376 (97.4) 
*

Global chi-square test for homogeneity.

Degree of organomegaly and size of mediastinal mass were determined as described in Materials and Methods.

Statistical methods.My+ BL and My+ TL patients were compared with their respective My BL and My TL controls for similarity of clinical, demographic, and laboratory features, as well as induction therapy outcome using global chi-square tests for homogeneity of proportions. Comparisons of antigen expression frequency distributions were performed using the Kruskal-Wallis nonparametric rank test.29 Most of the outcome analyses used life table methods and associated statistics. The primary endpoint was event-free survival (EFS) from the date of study entry. An event was defined as induction failure (no response to therapy or death during induction), leukemic relapse at any site, death during remission, or the development of a second malignant neoplasm, whichever occurred first. Patients not experiencing an event at the time of analysis were censored in the EFS analysis at the time of their last contact. Data analysis was performed in July 1996.

Life-table estimates were calculated by the Kaplan-Meier (KM) procedure, and the standard deviation (SD) of the life table estimate was obtained using Greenwood's formula.30 To indicate precision, the KM estimate of EFS and its SD were given for selected follow-up time points. An approximate 95% confidence interval can be obtained by using the life-table estimate ± 1.96 SDs. Life-table comparisons of EFS outcome pattern for patient groups used the log-rank statistic.31,32 Stratified log-rank tests were sometimes used to adjust for the possible modifying effects of other factors on the comparison of interest.32,33 P values for life-table comparisons are based on the pattern of outcome across the entire period of patient follow-up, although EFS estimates at specific time points may be given for comparative purposes. Estimates of the life-table relative hazard rate (RHR) for a particular event were calculated by the O/E method for log-rank analyses.34 

Table 3.

Presenting Features of Children With TL According to Myeloid Antigen Expression

VariableCategoryMy+ TL (N = 43)My TL (N = 184)P Value3-150
No.(%)No.(%)
Age (yr) <1 (2.3) (0.5) .003 
 1-9 18 (41.9) 126 (68.5)  
 ≥10 24 (55.8) 57 (31.0)  
WBC (×109/L) 1-19 13 (30.2) 51 (27.7) .93 
 20-49 (11.6) 24 (13.0)  
 ≥50 25 (58.1) 109 (59.2)  
Sex Male 34 (79.1) 133 (72.3) .47 
 Female (20.9) 51 (27.7)  
Race White 29 (67.4) 135 (73.8) .54 
 Black (16.3) 19 (10.4)  
 Other (16.3) 29 (15.8)  
Down syndrome Yes (0.0) (1.6) .92 
 No 43 (100.0) 180 (98.4)  
Liver Normal 28 (65.1) 54 (29.8) .12 
 Mod. enlarged3-151 13 (30.2) 108 (59.7)  
 Markedly enlarged (4.7) 19 (10.5)  
Spleen Normal 21 (48.8) 155 (29.9) .06 
 Mod. enlarged 16 (37.2) 93 (50.5)  
 Markedly enlarged (14.0) 36 (19.6)  
Lymph nodes Normal 18 (41.9) 44 (23.9) .05 
 Mod. enlarged 15 (34.9) 76 (41.3)  
 Markedly enlarged 10 (23.3) 64 (34.8)  
Mediastinal mass Absent 29 (67.4) 77 (41.8) .008 
 Small (11.6) 26 (14.1)  
 Large (20.9) 81 (44.0)  
Hemoglobin (g/dL) 1-7.9 12 (27.9) 50 (27.8) .02 
 8.0.-10.9 22 (51.2) 56 (31.1)  
 ≥11.0 (20.9) 74 (41.1)  
Platelets (×109/L) 1-49 10 (23.3) 74 (40.4) .11 
 50-149 19 (44.2) 63 (34.4)  
 ≥150 14 (32.6) 46 (25.1)  
CNS disease at diagnosis Yes (7.0) 16 (8.8) .93 
 No 40 (93.0) 165 (91.2)  
Karyotypic features 
Number Diploid (46) (47.1) 36 (43.9) .96 
 Hypodiploid (<46) (0.0) (2.4)  
 Pseudodiploid (46) (41.2) 33 (40.2)  
 Hyperdiploid (47-50) (11.8) 10 (12.2)  
 Hyperdiploid (>50) (0.0) (1.2)  
Aberrations Normal (47.1) 36 (43.9) .98 
 Abnormal (52.9) 46 (56.1) 
VariableCategoryMy+ TL (N = 43)My TL (N = 184)P Value3-150
No.(%)No.(%)
Age (yr) <1 (2.3) (0.5) .003 
 1-9 18 (41.9) 126 (68.5)  
 ≥10 24 (55.8) 57 (31.0)  
WBC (×109/L) 1-19 13 (30.2) 51 (27.7) .93 
 20-49 (11.6) 24 (13.0)  
 ≥50 25 (58.1) 109 (59.2)  
Sex Male 34 (79.1) 133 (72.3) .47 
 Female (20.9) 51 (27.7)  
Race White 29 (67.4) 135 (73.8) .54 
 Black (16.3) 19 (10.4)  
 Other (16.3) 29 (15.8)  
Down syndrome Yes (0.0) (1.6) .92 
 No 43 (100.0) 180 (98.4)  
Liver Normal 28 (65.1) 54 (29.8) .12 
 Mod. enlarged3-151 13 (30.2) 108 (59.7)  
 Markedly enlarged (4.7) 19 (10.5)  
Spleen Normal 21 (48.8) 155 (29.9) .06 
 Mod. enlarged 16 (37.2) 93 (50.5)  
 Markedly enlarged (14.0) 36 (19.6)  
Lymph nodes Normal 18 (41.9) 44 (23.9) .05 
 Mod. enlarged 15 (34.9) 76 (41.3)  
 Markedly enlarged 10 (23.3) 64 (34.8)  
Mediastinal mass Absent 29 (67.4) 77 (41.8) .008 
 Small (11.6) 26 (14.1)  
 Large (20.9) 81 (44.0)  
Hemoglobin (g/dL) 1-7.9 12 (27.9) 50 (27.8) .02 
 8.0.-10.9 22 (51.2) 56 (31.1)  
 ≥11.0 (20.9) 74 (41.1)  
Platelets (×109/L) 1-49 10 (23.3) 74 (40.4) .11 
 50-149 19 (44.2) 63 (34.4)  
 ≥150 14 (32.6) 46 (25.1)  
CNS disease at diagnosis Yes (7.0) 16 (8.8) .93 
 No 40 (93.0) 165 (91.2)  
Karyotypic features 
Number Diploid (46) (47.1) 36 (43.9) .96 
 Hypodiploid (<46) (0.0) (2.4)  
 Pseudodiploid (46) (41.2) 33 (40.2)  
 Hyperdiploid (47-50) (11.8) 10 (12.2)  
 Hyperdiploid (>50) (0.0) (1.2)  
Aberrations Normal (47.1) 36 (43.9) .98 
 Abnormal (52.9) 46 (56.1) 
F3-150

Global chi-square test for homogeneity.

F3-151

Degree of organomegaly and size of mediastinal mass were determined as described in Materials and Methods.

Immunophenotypic features of primary leukemic cells from children with My+ and My ALL.In accordance with the algorithm used for immunophenotypic classification, all BL patients showed high expression frequency for CD19 and all TL patients showed high expression frequency of CD7 (Table 1). Leukemic cells from BL patients were negative for T-lineage differentiation antigens and leukemic cells from TL patients were negative for B-lineage differentiation antigens (data not shown). The median expression frequencies of CD13 and CD33 were greater for My+ BL and My+ TL patients compared with My BL and My TL patients.

Fig. 1.

EFS of children with ALL according to BL immunophenotype. Percentages of 217 My+ BL (hatched line) and 1,113 My BL (solid line) patients achieving EFS during 6 years of follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Fig. 1.

EFS of children with ALL according to BL immunophenotype. Percentages of 217 My+ BL (hatched line) and 1,113 My BL (solid line) patients achieving EFS during 6 years of follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Close modal

Within the BL group, My+ and My patients had identical 88% median expression frequencies of CD10. Median expression frequencies for the CD34 and CD40 antigens were significantly higher in the My+ BL group than in the My BL group (P < .0001 for both comparisons). Similar immunophenotypic comparisons were performed for My+ TL and My TL patients (Table 1). Expression frequencies of CD2 and CD10 were similar for both groups. In contrast, the median expression frequency of CD5 was significantly lower for My+ TL patients compared with My TL patients (78% v 93%, P = .001). As was observed for BL patients, the median expression frequency of CD34 was significantly higher for the My+ TL patients compared with the My TL control group (60% v 8%, P = .0002).

Presenting features of children with My+ and My BL ALL.Clinical and laboratory features of My+ BL and My BL patients were compared by a global chi-square statistic (Table 2). WBC differed significantly between the two groups due to a higher percentage of My+ BL patients presenting with a low (<20,000/μL) WBC (68.7% v 58.1%; P = .006). The median WBC counts for My+ BL and My BL patients were 9,900 (range, 800 to 507,800) and 14,400 (range, 300 to 1,000,000), respectively. My+ BL patients were less likely than My BL patients to present with splenomegaly (47.0% v 57.2%; P = .0002) or lymphadenopathy (36.4% v 48.6%; P = .004). Platelet count also was significantly different (P = .01) between the two groups: My+ BL patients less often had low (<50,000/μL) and more often had high (≥150,000/μL) platelet counts at presentation. Centrally reviewed cytogenetic analysis was performed on leukemic cells from a subset of 71 My+ BL and 386 My BL patients. Within this subset, there were two significant differences between the groups. First, chromosome number differed significantly (P = .003) due both to the greater frequency of My+ BL patients presenting with a normal diploid karyotype (43.7% v 24.1%) and to the lower frequency of My+ BL patients presenting with high hyperdiploid (>50 chromosomes) karyotype (12.7% v 29.0%). Second, chromosomal aberrations were more frequent in the My BL group (75.9% v 53.6%, P = .001).

Fig. 2.

EFS of children with ALL according to TL immunophenotype. Percentages of 43 My+ TL (hatched line) and 184 My TL (solid line) patients achieving EFS during 5 years of follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Fig. 2.

EFS of children with ALL according to TL immunophenotype. Percentages of 43 My+ TL (hatched line) and 184 My TL (solid line) patients achieving EFS during 5 years of follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Close modal

Presenting features of children with My+ and My TL ALL.Clinical and laboratory features of My+ TL and My TL patients were compared in a similar manner (Table 3). Age distribution was significantly different (P = .003) for the My+ TL versus My TL groups largely due to a higher percentage of My+ TL patients (55.8% v 31.0%) presenting with ≥ 10 years of age. A higher percentage of My+ TL patients than My TL patients presented with a normal liver and spleen; however, these differences did not reach statistical significance. My+ TL patients were less likely than My TL patients to present with lymphadenopathy (58.2% v 76.1%, P = .05), a mediastinal mass (32.5% v 58.1%, P = .008), or high (≥11 g/dL) hemoglobin values (20.9% v 41.1%, P = .02). Cytogenetic data was available for only a small subset of patients (17 My+ TL and 82 My TL patients), and within this subset, there were no significant differences between the My+ TL and My TL patients.

Treatment outcomes for children with My+ and My ALL.Induction therapy outcomes were similar for My+ and My controls. At the end of induction chemotherapy, 98.6% of My+ BL patients and 98.3% of My BL patients achieved a remission (P = .97). Similarly, 97.6% of My+ TL patients and 96.6% of My TL patients achieved remission (P = .85). EFS outcomes of My+ BL and My+ TL patients were compared with those of My BL and TL patients using life-table methods, as described in Materials and Methods. Follow-up for event-free survivors ranged from 1 to 73 months (median, 32 months). My+ and My BL patients had similar outcomes (P = .32; Fig 1), with 4-year EFS estimates of 77.0% (SD = 4.0%) and 75.9% (SD = 1.8%), respectively. Similarly, the My+ TL and My TL groups had similar outcomes (P = .64; Fig 2 ), with 4-year EFS estimates of 72.7% (SD = 7.1%) and 70.1% (SD = 5.7%), respectively. The estimated RHR values for My+ BL versus My BL and My+ TL versus My TL were 0.83 and 1.18, respectively (Table 4). An overall RHR estimate of 0.89 (P = .49) was determined for My+ patients compared with My patients by a stratified analysis across lineage groups (Table 4).

Table 4.

RHR for My+ Versus My ALL Patients

BLTLCross-Strata
My+ BLMy BLMy+ TLMy TLMy+ ALLMy ALL
Observed events 35 209 11 41 46 250 
Expected events 40.81 203.19 9.68 42.32 50.49 245.51 
O/E 0.86 1.03 1.14 0.97 0.91 1.02 
RHR 0.83 
  1.18 
   0.89 
     
P value4-150 .32 
  .64 
   .49 
     
BLTLCross-Strata
My+ BLMy BLMy+ TLMy TLMy+ ALLMy ALL
Observed events 35 209 11 41 46 250 
Expected events 40.81 203.19 9.68 42.32 50.49 245.51 
O/E 0.86 1.03 1.14 0.97 0.91 1.02 
RHR 0.83 
  1.18 
   0.89 
     
P value4-150 .32 
  .64 
   .49 
     
F4-150

P values were calculated from a global chi-square statistic.

In addition, outcomes remained similar when BL patients were compared across NCI standard and poor risk group categories (P = .35 and P = .85, respectively; Fig 3). By this analysis, 4-year EFS estimates for My+ and My patients were 85.7% (SD = 3.9%) and 83.2% (SD = 2.1%), respectively, within the standard risk group, and 58.7% (SD = 9.1%) and 64.5% (SD = 3.2%), respectively, within the poor risk group. Estimates of RHR for My+ versus My patients in standard and poor-risk groups were 0.77 and 0.95, respectively. A stratified risk analysis was also performed to compare patients on CCG protocols with less intensive therapy (CCG 1881 and CCG 1891) with those on protocols with more intensive therapies (CCG 1882, CCG 1883, and CCG 1901). This analysis also showed similar outcome for My+ and My patients in low and high intensity treatment categories (P = .44 and P = .94, respectively; data not shown).

Fig. 3.

EFS of children with My+ and My BL ALL according to NCI risk classification. (Top) Standard risk: 138 My+ BL (hatched line) and 657 My BL (solid line) patients were followed for 6 years. (Bottom) Poor risk: 79 My+ BL (hatched line) and 456 My BL (solid line) were followed for 5 years. Percentages of patients achieving EFS during follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Fig. 3.

EFS of children with My+ and My BL ALL according to NCI risk classification. (Top) Standard risk: 138 My+ BL (hatched line) and 657 My BL (solid line) patients were followed for 6 years. (Bottom) Poor risk: 79 My+ BL (hatched line) and 456 My BL (solid line) were followed for 5 years. Percentages of patients achieving EFS during follow-up were calculated as described in Materials and Methods. The number of patients in each group remaining in follow-up at the indicated time points is shown in the inset.

Close modal

We have examined the clinical importance of myeloid antigen expression in a large cohort of children enrolled in risk-adjusted treatment protocols of the CCG. In general, children with My+ ALL compared with My ALL had similar or more favorable presenting features, including low WBC levels and normal karyotypes, as well as absence of splenomegaly, lymphadenopathy, and mediastinal mass. Importantly, we observed that remission induction rates, as well as EFS outcomes, were virtually identical for the My+ patients and My patients, demonstrating that myeloid antigen expression was not an adverse risk factor in this cohort.

Patients analyzed herein were defined according to coexpression of the myeloid differentiation antigens CD13 and CD33 together with either B (CD19) or T-(CD7) lymphoid–associated antigens. CD13, CD33, CD14, CD15, and CDw65 are the myeloid-associated antigens most frequently expressed on the surface of leukemic cells from ALL patients.35 Moreover, Drexler and Ludwig35 found that among ALL patients from numerous studies, similar percentages were positive for each of the individual myeloid antigens. Also, previous studies have documented the expression of various combinations of these myeloid-associated antigens in 5% to 20% of pediatric ALL patients,5 10-20 and differences in outcome do not appear to be related to the choice of antigens examined. Thus, analysis of CD13 and CD33 should be representative of overall myeloid antigen expression.

My+ ALL patients in the current study showed higher expression frequency of both CD34 and CD40 than My ALL patients. Similarly, Borowitz et al36 and Guyotat et al37 observed that in pediatric and adult patients, CD34 expression was correlated with myeloid antigen expression. Saeland et al38 reported that CD40 is also present on CD34+ immature myeloid progenitor cells, but is lost on interleukin-3 (IL-3) induced myeloid differentiation. CD34 is a 110 kD integral membrane protein thought to be expressed normally by immature hematopoietic progenitor cells,39,40 and CD40, a member of the nerve growth factor receptor superfamily, plays a role in proliferation and differentiation of normal B-lineage lymphoid cells.41-44 Therefore, expression of the CD34 and CD40 antigens by My+ ALL cells further supports the hypothesis that My+ ALL arises via transformation of an immature progenitor cell.

The clinical significance of myeloid antigen expression in children with ALL is controversial. In a single institution study involving 53 children with My+ ALL and 183 children with My ALL, Wiersma et al20 reported 3-year EFS estimates of 84% for My patients with WBC <50,000/μL, 57% for My patients with WBC ≥50,000/μL, 47% for My+ patients with WBC<50,000/μL, and 26% for My+ ALL patients with WBC ≥50,000/μL. These differences were statistically significant and multivariate analysis indicated that myeloid antigen expression was the most important predictor of a poor EFS outcome. Wiersma's study concurs with reports by Cantu Rajnoldi et al,14 Kurec et al,5 and Fink et al.15 Interestingly, these results also were consistent with preclinical observations that leukemic cells from My+ ALL patients were more resistant to glucocorticoid-induced killing than cells from My ALL patients.45 

Although the studies described above demonstrated a poor outcome associated with myeloid antigen expression in childhood ALL, numerous investigators have reported conflicting results. Bradstock et al13 and Mirro et al24 reported that myeloid antigen expression in pediatric ALL was not correlated with induction outcome, and other studies later showed no effect of myeloid antigen expression on treatment outcome.10,11,16,19 Likewise, Pui et al17 reported that myeloid antigen expression in 61 of 372 children with newly diagnosed ALL treated at the St. Jude Children's Hospital had no effect on either induction outcome or EFS. In a follow-up study, myeloid antigen expression again lacked prognostic significance in 25 children with My+ ALL.18 Subsequently, Pui et al46 reported that the estimated 3-year EFS estimates for 50 children with My+ ALL and 260 children with MyALL were 85% and 75%, respectively. The St Jude researchers concluded that myeloid antigen expression in childhood ALL is not associated with poor outcome if intensive chemotherapy regimens are used. Our results are generally consistent with these studies in showing that myeloid antigen expression does not correlate with poor outcome for children with ALL. In conclusion, this study provides new insight on the clinical significance of myeloid antigen expression in childhood ALL and shows that regardless of risk classification, ALL patients who are My+ have treatment outcomes similar to those who are My.

Supported in part by research grants including CCG Chairman's Grants No. CA-13539, CA-51425, CA-42633, CA-42111, CA-60437, and CA-27137 from the National Cancer Institute, National Institutes of Health, Bethesda, MD. F.M.U. is a Stohlman Scholar of the Leukemia Society of America.

Address reprint requests to Fatih M. Uckun, MD, Children's Cancer Group ALL Biology Reference Laboratory, Hughes Institute, 2657 Patton Rd, St Paul, MN 55113.

1
Pullen
DJ
Boyett
JM
Crist
WM
Falletta
JM
Roper
M
Dowell
B
Van Eys
J
Jackson
JF
Humphrey
GB
Metzgar
RS
Cooper
MD
Pediatric oncology group utilization of immunologic markers in the designation of acute lymphocytic leukemia subgroups: Influence on treatment response.
Ann NY Acad Sci
428
1984
26
2
Bene
MC
Castoldi
G
Knapp
W
Ludwig
WD
Matutes
E
Orfao
A
vant Veer MB
Proposals for the immunological classification of acute leukemias. European Group for the Immunological Characterization of Leukemias (EGIL).
Leukemia
9
1995
1783
3
Pui
CH
Behm
FG
Crist
WM
Clinical and biologic relevance of immunologic marker studies in childhood acute lymphoblastic leukemia.
Blood
82
1993
343
4
Sallan SE, Ritz J, Pesando J, Gelber R, O'Brien C, Hitchcock S, Coral F, Schlossman SF: Cell surface antigens: Prognostic implications in childhood acute lymphoblastic leukemia. Blood 55:395, 1980
5
Kurec
AS
Belair
P
Stefanu
C
Barrett
DM
Dubowy
RL
Davey
FR
Significance of aberrant immunophenotypes in childhood acute lymphoid leukemia.
Cancer
67
1991
3081
6
Ludwig
WD
Bartram
CR
Ritter
J
Raghavachar
A
Hiddemann
W
Heil
G
Harbott
J
Seibt
Jung H
Teichmann
JV
Riehm
H
Ambiguous phenotypes and genotypes in 16 children with acute leukemia as characterized by multiparameter analysis.
Blood
71
1988
1518
7
Champlin
R
Gale
RP
Acute lymphoblastic leukemia: Recent advances in biology and therapy [see comments].
Blood
73
1989
2051
8
Greaves
MF
Differentiation-linked leukemogenesis in lymphocytes.
Science
234
1986
697
9
Reinherz
EL
Kung
PC
Goldstein
G
Levey
RH
Schlossman
SF
Discrete stages of human intrathymic differentiation: Analysis of normal thymocytes and leukemic lymphoblasts of T-cell lineage.
Proc Natl Acad Sci USA
77
1980
1588
10
Ludwig
WD
Teichmann
JV
Sperling
C
Komischke
B
Ritter
J
Reiter
A
Odenwald
E
Sauter
S
Riehm
H
Incidence, clinical markers and prognostic significance of immunologic subtypes of acute lymphoblastic leukemia (ALL) in children: Experiences of the ALL-BFM 83 and 86 studies.
Klin Padiatr
202
1990
243
11
Ludwig
WD
Harbott
J
Bartram
CR
Komischke
B
Sperling
C
Teichmann
JV
Seibt
Jung H
Notter
M
Odenwald
E
Nehmer
A
Thiel
E
Riehm
H
Incidence and prognostic significance of immunophenotypic subgroups in childhood acute lymphoblastic leukemia: Experience of the BFM study 86.
Recent Results Cancer Res
131
1993
269
12
Ludwig WD, Reiter A, Loffler H, Gokbuget, Hoelzer D, Riehm H, Thiel E: Immunophenotypic features of childhood and adult acute lymphoblastic leukemia (ALL): Experience of the German Multicentre Trials ALL-BFM and GMALL. Leuk Lymphoma 1:71, 1994 (suppl 1)
13
Bradstock
KF
Kirk
J
Grimsley
PG
Kabral
A
Hughes
WG
Unusual immunophenotypes in acute leukaemias: Incidence and clinical correlations.
Br J Haematol
72
1989
512
14
Cantu
Rajnoldi A
Putti
C
Saitta
M
Granchi
D
Foa
R
Schiro
R
Castagni
M
Valeggio
C
Jankovic
M
Miniero
R
Paulucci
P
Basso
G
Co-expression of myeloid antigens in childhood acute lymphoblastic leukaemia: Relationship with the stage of differentiation and clinical significance [see comments].
Br J Haematol
79
1991
40
15
Fink
FM
Koller
U
Mayer
H
Haas
OA
Grumayer
Panzer ER
Urban
C
Dengg
K
Mutz
I
Tuchler
H
Gatterer
Menz I
Knapp
W
Gadner
H
Prognostic significance of myeloid-associated antigen expression on blast cells in children with acute lymphoblastic leukemia. The Austrian Pediatric Oncology Group.
Med Pediatr Oncol
21
1993
340
16
Hsu
PN
Tien
HF
Wang
CH
Chen
YC
Shen
MC
Lin
DT
Lin
KH
Liang
DC
Lin
KS
A subset of acute lymphoblastic leukemia with coexpression of myeloid antigens: Prevalence and clinical significance.
J Formosan Med Assoc
90
1991
225
17
Pui
CH
Behm
FG
Singh
B
Rivera
GK
Schell
MJ
Roberts
WM
Crist
WM
Mirro
J Jr
Myeloid-associated antigen expression lacks prognostic value in childhood acute lymphoblastic leukemia treated with intensive multiagent chemotherapy.
Blood
75
1990
198
18
Pui
CH
Raimondi
SC
Head
DR
Schell
MJ
Rivera
GK
Mirro
J Jr
Crist
WM
Behm
FG
Characterization of childhood acute leukemia with multiple myeloid and lymphoid markers at diagnosis and at relapse [see comments].
Blood
78
1991
1327
19
Urbano
Ispizua A
Matutes
E
Villamor
N
Ribera
JM
Feliu
E
Montserrat
E
Granena
A
Vives
Corrons JL
Rozman
C
Clinical significance of the presence of myeloid associated antigens in acute lymphoblastic leukaemia.
Br J Haematol
75
1990
202
20
Wiersma
SR
Ortega
J
Sobel
E
Weinberg
KI
Clinical importance of myeloid-antigen expression in acute lymphoblastic leukemia of childhood [see comments].
N Engl J Med
324
1991
800
21
Greaves
MF
Chan
LC
Furley
AJ
Watt
SM
Molgaard
HV
Lineage promiscuity in hemopoietic differentiation and leukemia.
Blood
67
1986
1
22
Smith
LJ
Curtis
JE
Messner
HA
Senn
JS
Furthmayr
H
McCulloch
EA
Lineage infidelity in acute leukemia.
Blood
61
1983
1138
23
McCulloch
EA
Smith
LJ
Alder
S
Cellular lineages in normal and leukemic hemopoiesis.
Prog Clin Biol Res
134
1983
229
24
Mirro
J
Zipf
TF
Pui
CH
Kitchingman
G
Williams
D
Melvin
S
Murphy
SB
Stass
S
Acute mixed lineage leukemia: Clinicopathologic correlations and prognostic significance.
Blood
66
1985
1115
25
Steinherz
PG
Siegel
SE
Bleyer
WA
Kersey
J
Chard
R Jr
Coccia
P
Leiken
S
Lukens
J
Neerhout
R
Nesbit
M
Miller
DR
Reaman
G
Sather
H
Hammond
D
Lymphomatous presentation of childhood acute lymphoblastic leukemia.
Cancer
68
1991
751
26
Smith
M
Arthur
D
Camitta
B
Carroll
AJ
Crist
W
Gaynon
P
Gelber
R
Heerema
N
Korn
EL
Link
M
Murphy
S
Pui
CH
Pullen
J
Reamon
G
Sallan
SE
Sather
H
Shuster
J
Simon
R
Trigg
M
Tubergen
D
Uckun
F
Ungerleider
R
Uniform approach to risk classification and treatment assignment for children with acute lymphoblastic leukemia [see comments].
J Clin Oncol
14
1996
18
27
Uckun
FM
Ledbetter
JA
Immunobiologic differences between normal and leukemic human B-cell precursors.
Proc Natl Acad Sci USA
85
1988
8603
28
Uckun
FM
Muraguchi
A
Ledbetter
JA
Kishimoto
T
O'Brien
RT
Roloff
JS
Gajl
Peczalska K
Provisor
A
Koller
B
Biphenotypic leukemic lymphocyte precursors in CD2+CD19+ acute lymphoblastic leukemia and their putative normal counterparts in human fetal hematopoietic tissues.
Blood
73
1989
1000
29
Hollander M, Wolfe D: Nonparametric Statistical Methods. New York, NY, Wiley, 1973
30
Kaplan
E
Meier
P
Nonparametric estimation from incomplete observations.
J Am Stat Assoc
53
1958
457
31
Mantel
N
Evaluation of survival data and two new rank order statistics arising in its consideration.
Cancer Chemother Rep
50
1966
163
32
Peto
R
Pike
MC
Armitage
P
Breslow
N
Cox
DR
Howard
SV
Mantel
N
McPherson
K
Peto
J
Smith
PG
Design and analysis of randomized clinical trials requiring prolonged observation of each patient, II. Analysis and examples.
Br J Cancer
35
1977
1
33
Breslow N: Comparison of survival curves, in Buyse M, Staquet M, Sylvester M (eds): Cancer Clinical Trials: Methods and Practice. Oxford, UK, Oxford, 1988, p 381
34
Breslow
N
Analysis of survival data under the proportional hazards model.
Int Stat Rev
43
1975
45
35
Drexler
HG
Ludwig
WD
Incidence and clinical relevance of myeloid antigen-positive acute lymphoblastic leukemia.
Recent Results Cancer Res
131
1993
53
36
Borowitz
MJ
Shuster
JJ
Civin
CI
Carroll
AJ
Look
AT
Behm
FG
Land
VJ
Pullen
DJ
Crist
WM
Prognostic significance of CD34 expression in childhood B-precursor acute lymphocytic leukemia: A Pediatric Oncology Group study.
J Clin Oncol
8
1990
1389
37
Guyotat
D
Campos
L
Shi
ZH
Charrin
C
Treille
D
Magaud
JP
Fiere
D
Myeloid surface antigen expression in adult acute lymphoblastic leukemia.
Leukemia
4
1990
664
38
Saeland
S
Duvert
V
Caux
C
Pandrau
D
Favre
C
Valle
A
Durand
I
Charbord
P
de Vries
J
Banchereau
J
Distribution of surface-membrane molecules on bone marrow and cord blood CD34+ hematopoietic cells.
Exp Hematol
20
1992
24
39
Greaves MF, Brown J, Molgaard HV, Spurr NK, Robertson D, Delia D, Sutherland DR: Molecular features of CD34: A hemopoietic progenitor cell-associated molecule. Leukemia 1:31, 1992 (suppl 6)
40
Simmons
DL
Satterthwaite
AB
Tenen
DG
Seed
B
Molecular cloning of a cDNA encoding CD34, a sialomucin of human hematopoietic stem cells.
J Immunol
148
1992
267
41
Banchereau
J
Bazan
F
Blanchard
D
Briere
F
Galizzi
JP
van Kooten
C
Liu
YJ
Rousset
F
Saeland
S
The CD40 antigen and its ligand.
Annu Rev Immunol
12
1994
881
42
Stamenkovic
I
Clark
EA
Seed
B
A B-lymphocyte activation molecule related to the nerve growth factor receptor and induced by cytokines in carcinomas.
EMBO J
8
1989
1403
43
Ledbetter
JA
Shu
G
Gallagher
M
Clark
EA
Augmentation of normal and malignant B cell proliferation by monoclonal antibody to the B cell-specific antigen Bp50 (Cdw40).
J Immunol
138
1987
788
44
Uckun
FM
Gajl
Peczalska K
Myers
DE
Jaszcz
W
Haissig
S
Ledbetter
JA
Temporal association of CD40 antigen expression with discrete stages of human B-cell ontogeny and the efficacy of anti-CD40 immunotoxins against clonogenic B-lineage acute lymphoblastic leukemia as well as B-lineage non-Hodgkin's lymphoma cells.
Blood
76
1990
2449
45
Kaspers
GJ
Kardos
G
Pieters
R
Van Zantwijk
CH
Klumper
E
Hahlen
K
de Waal
FC
van Wering
ER
Veerman
AJ
Different cellular drug resistance profiles in childhood lymphoblastic and non-lymphoblastic leukemia: A preliminary report.
Leukemia
8
1994
1224
46
Pui C, Schell M, Raimondi S, Head D, Rivera G, Crist W, Behm F: Myeloid antigen expression in childhood acute lymphoblastic leukemia. N Engl J Med 325:1378, 1991 (letter)
Sign in via your Institution