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Supplement
The following CALGB/Alliance institutions participated in this study and contributed at least 1%
of patents. For each of these institutions current principal investigators are listed.
Wake Forest University School of Medicine, Winston-Salem, NC: David D. Hurd; The Ohio State
University Medical Center, Columbus, OH: Clara D. Bloomfield (grant no. U10CA077658);
University of Iowa Hospitals, Iowa City, IA: Daniel A. Vaena; North Shore University Hospital,
Manhasset, NY: Daniel R. Budman (grant no. U10CA035279); Roswell Park Cancer Institute,
Buffalo, NY: Ellis G. Levine (grant no. U10CA059518); Duke University Medical Center, Durham,
NC: Jeffrey Crawford (grant no. U10CA047577); University of Chicago Medical Center, Chicago,
IL: Hedy L. Kindler (grant no. U10CA041287); Washington University School of Medicine, St.
Louis, MO: Nancy L. Bartlett (grant no. U10CA077440); University of Tennessee Cancer Center,
Memphis: Harvey B. Niell; Ft. Wayne Medical Oncology/Hematology, Ft. Wayne, IN: Sreenivasa
Nattam; University of Massachusetts Medical Center, Worcester, MA: William V. Walsh; Dana
Farber Cancer Institute, Boston, MA: Harold J. Burstein (grant no. U10CA032291); Eastern
Maine Medical Center, Bangor, ME: Thomas H. Openshaw; Vermont Cancer Center, Burlington,
VT: Steven M. Grunberg; University of North Carolina, Chapel Hill, NC: Thomas C. Shea (grant
no. U10CA047559); Dartmouth Medical School, Lebanon, NH: Konstantin Dragnev; Weill
Medical College of Cornell University, New York, NY: John Leonard; SUNY Upstate Medical
University, Syracuse, NY: Stephen L. Graziano; University of Maryland Cancer Center,
Baltimore, MD: Martin J. Edelman; Western Pennsylvania Hospital, Pittsburgh, PA: John Lister;
Mount Sinai School of Medicine, New York, NY: Lewis R. Silverman; Rhode Island Hospital,
Providence, RI: William Sikov; Moores University of California San Diego Cancer Center, San
Diego, CA: Barbara A. Parker; Christiana Care Health Services, Inc., Newark, DE: Stephen S.
Grubbs.
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Treatment
Patients with cytogenetically normal acute myeloid leukemia (CN-AML) younger than 60 years
were treated on Cancer and Leukemia Group B (CALGB) trial 19808. Older patients (60 years)
were treated on CALGB protocols 8525, 8923, 9420, 9720 and 10201. Patients enrolled on
CALGB 19808 (n=71) were randomly assigned to receive induction chemotherapy with
cytarabine, daunorubicin, and etoposide with or without PSC-833 (valspodar), a multidrug
resistance protein inhibitor.1 On achievement of complete remission (CR), patients were
assigned to intensification with high-dose cytarabine and etoposide for stem-cell mobilization
followed by myeloablative treatment with busulfan and etoposide supported by autologous
peripheral blood stem-cell transplantation. Older patients (≥60 years) were all treated with
cytarabine/daunorubicin-based induction therapy followed by cytarabine-based consolidation
therapy. Patients on CALGB 8525 (n=18) were treated with induction chemotherapy consisting
of cytarabine in combination with daunorubicin and were randomly assigned to consolidation
with different doses of cytarabine followed by maintenance treatment.2 Patients on CALGB 8923
(n=15) were treated with induction chemotherapy consisting of cytarabine in combination with
daunorubicin and were randomly assigned to receive postremission therapy with cytarabine
alone or in combination with mitoxantrone.3 Patients on CALGB 9420 (n=5) and 9720 (n=96)
received induction chemotherapy consisting of cytarabine in combination with daunorubicin and
etoposide, with (CALGB 9420) or with/without (CALGB 9720) the multidrug resistance protein
modulator PSC-833.4,5 Patients on CALGB 9420 received postremission therapy with cytarabine
(2 g/m2/d) alone, and patients on CALGB 9720 received a single cytarabine/daunorubicin
consolidation course and were then randomly assigned to low-dose recombinant interleukin-2
maintenance therapy or none.6 Patients on CALGB 10201 (n=65) received induction
chemotherapy consisting of cytarabine and daunorubicin, with or without the BCL2 antisense
oblimersen sodium. The consolidation regimen included two cycles of cytarabine (2 g/m 2/d) with
or without oblimersen.7
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Definition of clinical endpoints
Clinical endpoints were defined as follows8: CR required a bone marrow aspirate with cellularity
greater than 20% and maturation of all cell lines, less than 5% blasts and no Auer rods; in the
peripheral blood, an absolute neutrophil count of ≥1,500/µL, platelet count of >100,000/µL, and
no leukemic blasts; and no evidence of extramedullary leukemia, all of which had to persist for
at least 1 month.8 Relapse was defined by the presence of ≥5% bone marrow blasts, or
circulating leukemic blasts, or the development of extramedullary leukemia. Disease-free
survival (DFS) was measured from the date of CR until the date of relapse or death; patients
alive and in CR were censored at last follow-up. Overall survival (OS) was measured from the
date of study entry until the date of death, and patients alive at last follow-up were censored.
Statistical Analyses
The “optimal” cutpoint is defined as the value of the continuous MERTK variable that best
separates low and high risk patients with respect to DFS and OS using the log rank test.9
All clinical endpoint analyses were age group-adjusted (ie, <60 years vs ≥60 years) when we
considered all patients since there were treatment regimen or intensity differences in the
treatment protocols for these two age groups. Multivariable age group-adjusted logistic
regression models were generated for attainment of CR, and multivariable age group-adjusted
proportional hazards models were constructed for DFS and OS using a limited backwards
elimination procedure.9 Variables considered for model inclusion and evaluated in univariable
models were: GAS6 expression status, TYRO3/AXL dual receptor status, MERTK expression,
European LeukemiaNet (ELN) genetic category (based on the presence/absence of FLT3-ITD,
and NPM1 and CEBPA mutations), GAS6 mutation-by-TYRO3/AXL dual receptor status
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interaction, mutation status of the TET2, WT1, IDH1, IDH2, RUNX1, ASXL1, and DNMT3A
genes, FLT3-tyrosine kinase domain mutation status, MLL partial tandem duplication (MLL
PTD) status, age group (≥60 years vs <60 years, retained in all models), sex, race (white vs
nonwhite), white blood cell (WBC) count, hemoglobin and platelet counts. RUNX1 was not
considered for CR and DFS due to small sample sizes. Variables significant at α=.20 from the
univariable analyses were considered for multivariable analyses. For the time-to-event
endpoints, the proportional hazards assumption was checked for each variable individually. The
specific variables included in outcome modeling are: CR: GAS6 (+ vs –), ELN (Favorable vs
Intermediate-I), WT1 (mutated vs wild-type), ASXL1 (mutated vs wild-type), ERG (high vs low),
BAALC (high vs low), platelet counts (50x109/L increase), WBC count (50x109/L increase),
extramedullary involvement (present vs absent), age group (≥60 years vs <60 years); DFS:
GAS6 (+ vs –), TYRO3/AXL dual receptor status (+ vs –), ELN (Favorable vs Intermediate-I),
WT1 (mutated vs wild-type), MLL-PTD (present vs absent), DNMT3A (R882 mutated vs nonR882 mutated and wild-type), ERG (high vs low), BAALC (high vs low), age group (≥60 years vs
<60 years); OS: GAS6 (+ vs –), TYRO3/AXL dual receptor status (+ vs –), ELN (Favorable vs
Intermediate-I), WT1 (mutated vs wild-type), MLL-PTD (present vs absent), RUNX1 (mutated vs
wild-type), DNMT3A (R882 mutated vs non-R882 mutated and wild-type), ERG (high vs low),
BAALC (high vs low), WBC count (50x109/L increase), age group (≥60 years vs <60 years). Age
group was retained regardless of statistical significance.
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REFERENCES
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Kolitz JE, George SL, Marcucci G, Vij R, Powell BL, Allen SL et al. P-glycoprotein
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Lee EJ, George SL, Caligiuri M, Szatrowski TP, Powell BL, Lemke S et al. Parallel
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Marcucci G, Moser B, Blum W, Stock W, Wetzler M, Kolitz JE et al. A phase III
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Table S1. Clinical outcomes according to TYRO3/AXL/MERTK receptor tyrosine kinases expression status
in de novo cytogenetically normal AML patients
Variable
CR
DFS
OS
OR (95% CI)
P
HR (95% CI)
P
HR (95% CI)
P
TYRO3+ vs TYRO3–
1.30 (0.69-2.44)
0.41
1.72 (1.18-2.50)
0.005
1.56 (1.14-2.13)
0.005
AXL+ vs AXL–
0.75 (0.30-1.84)
0.52
1.04 (0.66-1.65)
0.86
0.82 (0.54-1.25)
0.36
MERTK+ vs MERTK–
1.09 (0.61-1.92)
0.78
0.88 (0.63-1.22)
0.44
0.89 (0.68-1.17)
0.41
Abbreviations: CR, complete remission, DFS, disease-free survival, OS, overall survival; OR, odds ratio; CI, confidence interval; HR, hazard ratio.
All clinical endpoints are age group adjusted.
An odds ratio greater than (less than) 1.0 means a higher (lower) CR rate for patients in the first group compared to patients in the second group.
A hazard ratio greater than 1 (less than 1) corresponds to a higher (lower) risk of an event for patients in the first group compared to patients in
the second group.
The median follow-up for those patients alive is 7.2 years, range: 2.3-11.6 years
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Table S2. Genes differentially expressed between GAS6+ and GAS6- CN-AML patients
(see separate Excel file)