Combination of KIT and FLT3-ITD mutation status with minimal residual disease levels guides treatment strategy for adult patients with inv(16) acute myeloid leukemia in first complete remission.

Although several studies have investigated the benefits of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for patients with inv (16) acute myeloid leukemia (AML) in first complete remission (CR1) individually stratified by KIT or FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) mutation status or minimal residual disease (MRD) levels, evaluation based on the combination of mutation status and MRD levels remains absent. This study included 157 adult patients with inv (16) AML who were consecutively diagnosed and receiving treatment at our center. A total of 50 (31.6%) patients had KIT mutations (KITMU ), and the risk of relapse was significantly higher in patients with KITMU than in patients with KITWT (p < 0.001). A total of 12 patients (7.6%) had FLT3-ITD, and FLT3-ITD+ tended to be related to a higher risk of relapse (p = 0.14). KITMU , FLT3-ITD and MRD3-H (beta subunit of core binding factor-myosin heavy chain 11 levels >0.2% after course 2 of consolidation therapy) were independent adverse prognostic factors for relapse with patients who received allo-HSCT at CR1 were censored at the time of transplantation. After combination, patients were categorized into molecularly defined high-risk (M-HR; KITMU or FLT3-ITD+ with MRD3-H; n = 30), low-risk (M-LR; KITWT and FLT3-ITD- with MRD3-L; n = 45) and intermediate-risk (M-IR; others; n = 70) groups. For the M-HR group, allo-HSCT significantly improved both cumulative incidence of relapse cumulative incidence of relapse (CIR) and overall survival (OS) (11.1% vs. 92.6%, p < 0.001; 90.0% vs. 34.1%, p = 0.019). For the M-IR group, allo-HSCT significantly improved CIR but did not affect OS (14.1% vs. 62.2%, p = 0.0004; 73.3% vs. 68.3%, p = 0.43). For the M-LR group, allo-HSCT had no significant effect on both CIR and OS (0% vs. 35.1%, p = 0.31; 100% vs. 78.8%, p = 0.22). Therefore, the combination of KIT and FLT3-ITD mutation status with MRD levels may identify inv (16) AML patients with high-risk who can benefit from allo-HSCT in CR1.

Low EVI1 expression at diagnosis predicted poor outcomes in pediatric Ph-negative B cell precursor acute lymphoblastic leukemia patients.

Abnormally high ecotropic viral integration site 1 (EVI1) expression has been recognized as a poor prognostic factor in acute myeloid leukemia patients. However, its prognostic impact in B cell precursor acute lymphoblastic leukemia (BCP-ALL) remains unknown. A total of 176 pediatric Ph-negative BCP-ALL patients who received at least 1 course of chemotherapy and received chemotherapy only during follow-up were retrospectively tested for EVI1 transcript levels by real-time quantitative PCR at diagnosis, and survival analysis was performed. Clinical and EVI1 expression data of 129 pediatric BCP-ALL patients were downloaded from therapeutically applicable research to generate effective treatments (TARGET) database for validation. In our cohort, the median EVI1 transcript level was 0.33% (range, 0.0068-136.2%), and 0.10% was determined to be the optimal cutoff value for patient grouping by receiver operating characteristic curve analysis. Low EVI1 expression (<0.10%) was significantly related to lower 5-year relapse-free survival (RFS) and overall survival (OS) rates (P = 0.017 and 0.018, respectively). Multivariate analysis showed that EVI1 expression <0.10% was an independent adverse prognostic factor for RFS and OS. TARGET data showed that low EVI1 expression tended to be related to a lower 5-year OS rate (P = 0.066). In conclusion, low EVI1 expression at diagnosis could predict poor outcomes in pediatric Ph-negative BCP-ALL patients receiving chemotherapy.Supplemental data for this article is available online at https://doi.org/10.1080/08880018.2021.1939818 .

Prognostic significance of TIM-3 expression pattern at diagnosis in patients with t(8;21) acute myeloid leukemia.

Acute myeloid leukemia (AML) with t(8;21) is a heterogeneous disease and needs to be stratified. Both, cancer cells and immune cells participate in tumor initiation, growth and progression and might affect clinical outcomes. TIM-3 (T cell immunoglobulin and mucin domain-containing protein 3), an immune checkpoint molecule, is expressed not only on immune cells but also on leukemic stem cells (LSCs) in AML. This prompted us to investigate the prognostic significance of TIM-3 in t(8;21) AML. A total of 47 t(8;21) AML patients were tested for TIM-3 expression by multi-parameter flow cytometry at diagnosis. 35 of these, who received chemotherapy alone or along with allogeneic hematopoietic stem cell transplantation were followed up. The expression pattern of TIM-3 on T-cells and NK (natural killer) cells as a whole (T + NK) and LSCs were evaluated independently. High percentage of T + NK - TIM-3+ and CD34+CD38-TIM-3+ cells were significantly associated with a high 2-year cumulative incidence of relapse (CIR) (p = 0.028, 0.016). Further, concurrent high frequencies of T + NK-TIM-3+ and CD34+CD38-TIM-3+ cells at diagnosis were significantly associated with a high 2-year CIR (p < 0.0001) and this together with c-KIT D816 mutation were the independent adverse prognostic factors for relapse (hazard ratio (HR)=2.5, [95% confidence interval (CI), 1.1-6.0], p = 0.04; HR = 46.5, [95% CI, 2.7-811.5], p = 0.009). In conclusion, the expression pattern of TIM-3 on both T and NK cells and LSCs at diagnosis had prognostic significance in t (8;21) AML.

The Prognostic Significance of ZNF384 Fusions in Adult Ph-Negative B-Cell Precursor Acute Lymphoblastic Leukemia: A Comprehensive Cohort Study From a Single Chinese Center.

Novel recurrent fusion gene types such as zinc finger protein 384 (ZNF384) fusions have been identified in B-cell precursor acute lymphoblastic leukemia (BCP-ALL) with the application of next-generation sequencing technologies. However, the comprehensive large-scale clinical cohort study for clarifying their prognostic significance remains scarce to date. A total of 242 consecutive adult Ph-negative BCP-ALL patients treated in our institute were retrospectively screened ZNF384 fusions at diagnosis by multiplex real time quantitative PCR. ZNF384 fusions were identified in 47 patients (19.4%) and all belonged to B-other ALL (having no high hyperdiploid karyotype, BCR-ABL1, TCF3-PBX1, ETV6-RUNX1, or MLL rearrangement). In the whole cohort, patients with ZNF384 fusions had significantly higher 3-year relapse-free-survival (RFS) and tended to have a higher 3-year overall survival (OS) than those with no ZNF384 fusions (80.1% vs. 52.5%, P = 0.013; 67.6% vs. 54.0%, P = 0.10). For patients receiving chemotherapy alone and received allogeneic-hematologic stem cell transplantation (allo-HSCT) were censored at the time of transplantation, patients with ZNF384 fusions had both similar RFS and similar OS to B-other ALL patients with no ZNF384 fusions (RFS: P =0.94 and 0.30; OS: P =0.94 and 0.51). For patients receiving transplantation, those with ZNF384 fusions had significantly higher 3-year RFS than B-other ALL patients with no ZNF384 fusions and their OS were similar (P = 0.022 and 0.24). Only two of 31 patients with ZNF384 fusions and receiving allo-HSCT relapsed, individually occurred 66.8 and 69.8 months after transplantation. Therefore, ZNF384 fusion is common in adult BCP-ALL, which may define a new group from BCP-ALL containing no classical fusion transcript with better prognosis through receiving allo-HSCT.

Development of a poor-prognostic-mutations derived immune prognostic model for acute myeloid leukemia.

Leukemia cell-intrinsic somatic mutations and cytogenetic abnormalities have been used to define risk categories in acute myeloid leukemia (AML). In addition, since the immune microenvironment might influence prognosis and somatic mutations have been demonstrated to modulate the immune microenvironment in AML, there is need for developing and evaluating an immune prognostic model (IPM) derived from mutations associated with poor prognosis. Based on AML cases with intermediate and adverse-cytogenetic risk in the Cancer Genome Atlas (TCGA) database, 64 immune-related differentially expressed genes (DEGs) among patients with RUNX1, TP53, or ASXL1 mutations and patients without these mutations were identified. After Cox proportional hazards analysis, an IPM composed of PYCARD and PEAR1 genes was constructed. IPM defined high-risk (IPM-HR) independently predicted lower 2-year overall survival (OS) rates in both patients with intermediate and adverse-cytogenetic risks and non-M3 patients in the TCGA AML cohort. The poor prognostic impact of IPM-HR on OS was further validated by GSE71014, 37642, and 10358 downloaded from the Gene Expression Omnibus (GEO) database. Furthermore, IPM-HR was remarkably associated with higher proportions of CD8+ T cells and regulatory T cells (Tregs), lower proportions of eosinophils, and higher expression of the checkpoint molecules CTLA-4, PD-1, and LAG3 in the TCGA non-M3 AML cohort. In summary, we developed and validated an IPM derived from mutations related with poor prognosis in AML, which would provide new biomarkers for patient stratification and personalized immunotherapy.

[The Predict Significance of ALDH Activity to the Relapse of t(8;21) Acute Myeloid Leukemia Patients at Complete Remission].

OBJECTIVE: To investigate the predict significance of the high aldehyde dehydrogenase activity (ALDH+) propitiation to the relapse of t(8;21) acute myeloid leukemia(AML) patients before and after treatment. METHODS: Bone marrow samples of 23 t(8;21) AML patients diagnosis and achieved complete remission in our hospital from April 2015 to June 2016 were collected, then flow cytometry method was used to detect the activity of ALDH, relationship between it and relapse was analyzed. RESULTS: All the patients were followed up for a median of 32 (2-52) months. The median percentage of CD34+CD38- and CD34+CD38-ALDH+ cells among nucleated cells were 2.7 (0.36-35.7)% and 0.017 (0.0013-0.62)% at diagnosis, respectively. Among the bone marrow samples in patients achieved CR, the median percentage of CD34+CD38- cells was 0.035 (0.0064-0.66)%, and it was significantly decreased as compared with the percentage at diagnosis (P<0.001); The median percentage of CD34+CD38-ALDH+ cells was 0.014 (0.0019-0.24)%, and it showed no different as compared with the percentage at diagnosis (P=0.45). Survival analysis showed that patients with higher percentage of CD34+CD38- and CD34+CD38-ALDH+ cells at diagnosis tended to the lower 3-year relapse-free survival (RFS) (P=0.27 and 0.21). Furthermore, patients with higher percentage of CD34+CD38- and CD34+CD38-ALDH+ cells when achieved CR had a significant lower 3-year RFS rates (P=0.010 and 0.0044) as compared with those with lower percentage of CD34+CD38- and CD34+CD38-ALDH+ cells. Multivariate analysis showed that higher percentage of CD34+CD38-ALDH+ cells when achieved CR was an independent factor affecting RFS of the patients. CONCLUSION: The percentage of CD34+CD38-ALDH+ cells at CR in t(8;21) AML patients could predicts relapse, and had more profound predictive significance for relapse than the percentage of CD34+CD38- cells.

High PRDM16 expression predicts poor outcomes in adult acute myeloid leukemia patients with intermediate cytogenetic risk: a comprehensive cohort study from a single Chinese center.

Acute myeloid leukemia with intermediate cytogenetic risk (ICR-AML) needs to be stratified and abnormal gene expression might be prognostic. PR/SET domain 16 (PRDM16) transcript levels were assessed in 267 consecutive adult ICR-AML patients at diagnosis by real-time quantitative PCR. 38.2% patients had PRDM16 transcript levels higher than the upper limit of normal bone marrow samples. Through ROC curve analysis and comparison of relapse-free survival (RFS), the optimal cutoff value of PRDM16 transcript levels was identified to group patients into high expression (PRDM16-H, 21.3%) and low expression (PRDM16-L). PRDM16-H was significantly associated with lower 4-year RFS and overall survival (OS) rates in the entire cohort, patients with normal karyotypes, FLT3-ITD (-) and NPM1 mutation (+)/FLT3-ITD (-) patients (all p < .05). Multivariate analysis showed that PRDM16-H was an independent adverse prognostic factor for RFS and OS in the entire cohort. Therefore, high PRDM16 expression at diagnosis predicts poor outcomes in adult ICR-AML patients.

Characterization of somatic mutation-associated microenvironment signatures in acute myeloid leukemia patients based on TCGA analysis.

Recurrent genetic mutations occur in acute myeloid leukemia (AML) and have been incorporated into risk stratification to predict the prognoses of AML patients. The bone marrow microenvironment plays a critical role in the development and progression of AML. However, the characteristics of the genetic mutation-associated microenvironment have not been comprehensively identified to date. In this study, we obtained the gene expression profiles of 173 AML patients from The Cancer Genome Atlas (TCGA) database and calculated their immune and stromal scores by applying the ESTIMATE algorithm. Immune scores were significantly associated with OS and cytogenetic risk. Next, we categorized the intermediate and poor cytogenetic risk patients into individual-mutation and wild-type groups according to RUNX1, ASXL1, TP53, FLT3-ITD, NPM1 and biallelic CEBPA mutation status. The relationships between the immune microenvironment and each genetic mutation were investigated by identifying differentially expressed genes (DEGs) and conducting functional enrichment analyses of them. Significant immune- and stromal-relevant DEGs associated with each mutation were identified, and most of the DEGs (from the FLT3-ITD, NPM1 and biallelic CEBPA mutation groups) were validated in the GSE14468 cohort downloaded from the Gene Expression Omnibus (GEO) database. In summary, we identified key immune- and stromal-relevant gene signatures associated with genetic mutations in AML, which may provide new biomarkers for risk stratification and personalized immunotherapy.

Both Methylation and Copy Number Variation Participated in the Varied Expression of PRAME in Multiple Myeloma.

PURPOSE: The cancer-testis antigen, which is a preferentially expressed antigen of melanoma (PRAME), is an ideal target for immunotherapy and cancer vaccines. Since the expression of this antigen is relevant to therapy responses, the heterogeneity in its expression and the underlying mechanism need to be investigated. PATIENTS AND METHODS: Plasma cell sorting was performed in 48 newly diagnosed multiple myeloma (MM) patients. Real-time quantitative PCR was performed to examine the PRAME transcript levels and gene copy numbers. Bisulfate clone sequencing of the PRAME promoter and exon 1b regions was performed in 4 patients. Quantitative methylation-specific PCR of the +287 CpG site was performed for all patients. The human MM cell lines RPMI8226, LP-1 and MOLP-2 were treated with 5-azacytidine. RESULTS: The median PRAME transcript level was 3.1% (range: 0-298.3%) in the plasma cells sorted from the 48 MM patients. Eleven (22.9%) and 37 (77.1%) patients were individually categorized into the PRAME low- and high-expression groups according to the cut-off value of 0.05%. The methylation ratios of the promoter and the 3' region of exon 1b region were both negatively related to the transcript levels. The degrees of methylation at the +287 CpG site were significantly negatively related to the transcript levels in all 48 patients (r=-0.44, P=0.0018), and those in the high-expression group (r=-0.69, P<0.0001) but not those in the low-expression group (r=-0.27, P=0.43). All 5 patients with homozygous deletions were categorized into the low-expression group. There were no significant differences in the PRAME transcript levels between the hemizygous deletion (n=8) and no deletion (n=35) groups (P=0.40). Furthermore, the PRAME transcript levels significantly increased in the MM cell lines after treatment with 5-azacytidine. CONCLUSION: Both methylation and copy number variation may participate in the regulation of PRAME expression in MM; in patients with no homozygous deletion, PRAME expression is mainly controlled by methylation, and a proportion of fairly low expression is caused by homozygous deletion.

High aldehyde dehydrogenase activity at diagnosis predicts relapse in patients with t(8;21) acute myeloid leukemia.

Acute myeloid leukemia (AML) with t(8;21) is a heterogeneous disease. Although the detection of minimal residual disease (MRD), which is indicated by RUNX1-RUNX1T1 transcript levels, plays a key role in directing treatment, risk stratification needs to be improved, and other markers need to be assessed. A total of 66 t(8;21) AML patients were tested for aldehyde dehydrogenase (ALDH) activity by flow cytometry at diagnosis, and 52 patients were followed up for a median of 20 (1-34) months. The median percentage of CD34+ALDH+, CD34+CD38-ALDH+, and CD34+CD38+ALDH+ cells among nucleated cells were 0.028%, 0.012%, and 0.0070%, respectively. The CD34+ALDH+-H, CD34+CD38-ALDH+-H, and CD34+CD38+ALDH+-H statuses (the percentage of cells that were higher than the individual cutoffs) were all significantly associated with a lower 2-year relapse-free survival (RFS) rate in both the whole cohort and adult patients (P = .015, .016, and .049; P = .014, .018, and .032). Patients with < 3-log reduction in the RUNX1-RUNX1T1 transcript level after the second consolidation therapy (defined as MRD-H) had a significantly lower 2-year RFS rate than patients with ≥ 3-log reduction (MRD-L) (P = .017). The CD34+ALDH+ status at diagnosis was then combined with the MRD status. CD34+ALDH+-L/MRD-H patients had similar 2-year RFS rates to both CD34+ALDH+-L/MRD-L and CD34+ALDH+-H/MRD-L patients (P = .50 and 1.0); and CD34+ALDH+-H/MRD-H patients had significantly lower 2-year RFS rate compared with CD34+ALDH+-L and/or MRD-L patients (P < .0001). Multivariate analysis showed that CD34+ALDH+-H/MRD-H was an independent adverse prognostic factor for relapse. In conclusion, ALDH status at diagnosis may improve MRD-based risk stratification in t(8;21) AML, and concurrent high levels of CD34+ALDH+ at diagnosis and MRD predict relapse.