Patient characteristics
From February 2011 to February 2019, non-DS-AMKL was diagnosed in 40 patients at our institution. The last follow-up was November 2019, and the median follow-up period was 16.1 months (range, 4.6 to 71.8). Patients with AMKL frequently presented with a long disease history. The median interval to diagnosis at our institution was 1 month (range, 0-12 months). Additionally, there were no patients with initial CNS involvement in our cohort. Patient characteristics are summarized in Table 2.
Cytogenetics analysis
Cytogenetic data were available for 37 (92.3%) patients: hypodiploid (n=0), normal karyotype (n=9, 24.3%), pseudodiploid (n=8, 21.6%), 47 to 50 chromosomes (n=12, 32.4%), and >50 chromosomes (n=7, 18.9%). Among 28 patients with chromosomal aberrations, 5 (17.9%) had numerical aberrations only, 6(21.4%) had structural aberrations only, 17 (60.7%) had both, and 13 (46.4%) had complex karyotypes (three or more independent abnormalities including at least one structural abnormality). Chromosome gain such as +21, +8, +19 occurred in 13(35.1%), 12(32.4%), and 8(21.6%) patients, respectively. Three patients (8.1%) had 11q23 aberrations and only 1 patient had t (1;22) (p13; q13). Monosomy 7/7q- and -5/5q- were found in 3(8.1%) and 2(5.4%) patients, respectively.
Molecular analysis
Sequencing of the hematopoietic transcription factor GATA1 was performed on 22 patients. Four patients (18.2 %) were positive for a GATA1-mutation. None of these children was previously diagnosed with DS or had a history of TMD during the neonatal period. Interestingly, the leukemic blasts of all four patients showed CD7 surface marker expression, a common feature of DS-AMKL. Three patients were treated by chemotherapy alone and are in continuous CR. Only one child received HSCT due to the high level of MRD after induction Ⅰ. Each child with GATA1-mutation was maintained in continuous CR.
Rearrangements of the MLL gene on 11q23 via RT-PCR detection were detected in 4(10.0%) patients, and one of them was not detected by G-banding. The MLL translocation in this group was MLL-AF4 (n=1), MLL-AF9 (n=2) and MLL-PTD (n=1).
WT1 and EVI1 overexpression were identified in 33 of 37 (89.2%) and 17 of 29 (58.6%) patients, respectively.
Early treatment response
Two of the 40 patients transferred to another hospital after induction Ⅰ without evaluation of the BM status. One and two patients abandoned treatment with no CR at the end of induction Ⅰ and Ⅱ, respectively. Of the remaining patients, 22 (57.9%) and 30 (81.1%) were in CR after induction Ⅰ and Ⅱ, respectively. Twenty-six (68.4%) had an MRD < 1% and 12 (31.6%) had an MRD > 1% after induction Ⅰ (Table 2).
Outcomes
Ultimately, 35 patients were included for survival analysis. The transplant and nontransplant cohorts comprised of 26 and 9 patients, respectively. Of the 26 patients in the transplant cohort, 16 had haplo-HSCT, 7 had HLA-MSDT (including 3 had a 6/6 HLA-MSDT), 1 had an HLA-MUDT, and the other 2 had a UCB donor (1 with a 6/6 HLA-matched donor, 1 with a 4/6 HLA-matched donor). Twenty-three patients opted for HSCT at CR1. The remaining 3 patients underwent transplantation without achieving CR after relapse for further salvage treatment and just one of them achieved CR2 for only 2 months after HSCT. Besides the 3 patients, 7 patients relapsed ultimately after transplantation and the median time between HSCT and relapse was 3 months (range , 0-16 months).
The estimated 2-year probability of OS was 41±13% among 35 patients, 42±15% in the transplant cohort, and 41±17% in the chemotherapy cohort. The estimated 2-year probability of EFS was 41±10% in all patients, 49±12% in the transplant cohort, and 21±17% in the chemotherapy cohort. (Fig. 2 and 3)
Factors associated with long-term survival
The patients' clinical characteristics, including gender, age, WBC count at diagnosis, did not affect survival. Analysis of cytogenetic features for OS demonstrated that patients with +21 or hyperdiploid (>50 chromosomes) had significantly better OS than those in other cytogenetic subgroups (Plog-rank=0.048 and Plog-rank=0.040, respectively).
We also compared the response to induction therapy and survival outcomes between EVI1 high patients and patients with low/undetectable EVI1. EVI1 high patients had a CR rate of 57.1% after induction Ⅰ as compared to 90.9% for the remaining patients (P =0.053). Besides, patients in the EVI1 high cohort tended to have a lower rates of 2-year OS (54±14% vs 89±10%, Plog-rank=0.089) and EFS (54±14% vs 79±13%, Plog-rank=0.179); but, the data were not statistically significant. However, EVI1 high patients had a better OS in transplant cohort than in chemotherapy cohort (Plog-rank=0.009).
Early treatment response also had an impact on the survival rates of non-DS-AMKL patients. The poor responders had an estimated 2-year OS of only 8±11 % and 2-year EFS of 18±12 %, while good responders (<5 % BM blasts after induction Ⅰ) showed a 2-year OS of 74±12 % (Plog-rank=0.023) and 2-year EFS of 60±14 % (Plog-rank=0.003). Besides, patients with MRD < 1% tended to have a favorable OS and EFS compared to those with a high level of MRD at the end of induction Ⅰ (Table 3).
Next, we assessed the therapeutic effect of HSCT. The baseline characteristics were similar between the nontransplant and transplant cohorts (Table 2). Patients who did not receive HSCT and had an event before the median time until transplantation (4 months) were excluded. The estimated 2-year OS and EFS were significantly higher in the transplant cohort than in the chemotherapy cohort (Plog-rank=0.042 and 0.044, respectively).
In the multivariate analysis for 2-year OS and EFS, which included hyperdiploid, +21, early treatment response as well as post-remission therapy as risk factors, only post-remission therapy (transplant vs chemotherapy; RR=11.192; 95 % CI, 2.045-61.241; P=0.005 for OS and RR=5.400; 95 % CI, 1.635-17.832; P=0.006 for EFS, respectively) had independent prognostic significance. For poor early treatment response, which was statistically significant in the univariate analysis, multivariate analysis indicated a trend towards poor prognosis (Table 4).