Acute Myeloid Leukemia with the t(8;21) Translocation: Clinical Consequences and Biological Implications

The t(8;21) abnormality occurs in a minority of acute myeloid leukemia (AML) patients. The translocation results in an in-frame fusion of two genes, resulting in a fusion protein of one N-terminal domain from the AML1 gene and four C-terminal domains from the ETO gene. This protein has multiple effects on the regulation of the proliferation, the differentiation, and the viability of leukemic cells. The translocation can be detected as the only genetic abnormality or as part of more complex abnormalities. If t(8;21) is detected in a patient with bone marrow pathology, the diagnosis AML can be made based on this abnormality alone. t(8;21) is usually associated with a good prognosis. Whether the detection of the fusion gene can be used for evaluation of minimal residual disease and risk of leukemia relapse remains to be clarified. To conclude, detection of t(8;21) is essential for optimal handling of these patients as it has both diagnostic, prognostic, and therapeutic implications.

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Immunophenotyping Investigation of Minimal Residual Disease Is a Useful Approach for Predicting Relapse in Acute Myeloid Leukemia Patients

Blood ◽

10.1182/blood.v90.6.2465 ◽

1997 ◽

Vol 90 (6) ◽

pp. 2465-2470 ◽

Cited By ~ 177

Author(s):

J.F. San Miguel ◽

A. Martı́nez ◽

A. Macedo ◽

M.B. Vidriales ◽

C. López-Berges ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Multidrug Resistance ◽

Minimal Residual Disease ◽

Myeloid Leukemia ◽

Residual Disease ◽

Leukemic Cells ◽

Rhodamine 123 ◽

Minimal Residual ◽

Acute Myeloid

Abstract A high complete remission rate is currently achieved in patients with acute myeloid leukemia (AML). However, many patients eventually relapse due to the persistence of low numbers of residual leukemic cells that are undetectable by conventional cytomorphologic criteria (minimal residual disease [MRD]). Using immunophenotypic multiparametric flow cytometry, we have investigated in sequential studies (diagnosis and follow-up) the impact of MRD detection on the outcome of 53 AML patients that had achieved morphologic remission with standard AML protocols and displayed at diagnosis an aberrant phenotype. Patients were studied at diagnosis with a panel of 35 monoclonal antibodies in triple staining combinations for detection of aberrant or uncommon phenotypic features. According to these features, a patient's probe was custom-built at diagnosis for the identification of possible residual leukemic cells during follow-up. The level of MRD at the end of induction and intensification therapy correlated with the number of relapses and relapse-free survival (RFS). Thus, patients with more than 5 × 10−3 residual cells (5 residual cells among 1,000 normal bone marrow [BM] cells) identified as leukemic by immunophenotyping in the first remission BM showed a significant higher rate of relapse (67% v 20% for patients with less than 5 × 10−3 residual cells; P = .002) and a lower median RFS (17 months v not reached; P = .01). At the end of intensification, with a cut-off value of 2 × 10−3 leukemic cells, AML patients also separated into two distinct groups with relapse rates of 69% versus 32% (P = .02), respectively, and median RFS of 16 months versus not reached (P = .04). In addition, overall survival was also significantly related to the level of residual cells in the marrow obtained at the end of induction and particularly after intensification therapy (P = .008). Furthermore, we have explored whether residual disease was related with the functional expression of multidrug resistance (MDR-1) at diagnosis as assessed by the rhodamine-123 assay. Patients with ≥5 × 10−3 residual leukemic cells at the end of induction therapy had a significantly higher rhodamine-123 efflux (mean, 56% ± 24%) than those with less than 5 × 10−3 residual cells (mean, 32% ± 31%; P = .04). Finally, multivariate analysis showed that the number of residual cells at the end of induction or intensification therapy was the most important prognostic factor for prediction of RFS. Overall, our results show that immunophenotypical investigation of MRD strongly predicts outcome in patients with AML and that the number of residual leukemic cells correlates with multidrug resistance.

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Ezh2 augments leukemogenicity by reinforcing differentiation blockage in acute myeloid leukemia

Blood ◽

10.1182/blood-2011-11-394932 ◽

2012 ◽

Vol 120 (5) ◽

pp. 1107-1117 ◽

Cited By ~ 122

Author(s):

Satomi Tanaka ◽

Satoru Miyagi ◽

Goro Sashida ◽

Tetsuhiro Chiba ◽

Jin Yuan ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Target Genes ◽

Fusion Gene ◽

Hematologic Malignancies ◽

Leukemic Cells ◽

Polycomb Repressive Complex 2 ◽

Acute Myeloid

Abstract EZH2, a catalytic component of the polycomb repressive complex 2, trimethylates histone H3 at lysine 27 (H3K27) to repress the transcription of target genes. Although EZH2 is overexpressed in various cancers, including some hematologic malignancies, the role of EZH2 in acute myeloid leukemia (AML) has yet to be examined in vivo. In the present study, we transformed granulocyte macrophage progenitors from Cre-ERT;Ezh2flox/flox mice with the MLL-AF9 leukemic fusion gene to analyze the function of Ezh2 in AML. Deletion of Ezh2 in transformed granulocyte macrophage progenitors compromised growth severely in vitro and attenuated the progression of AML significantly in vivo. Ezh2-deficient leukemic cells developed into a chronic myelomonocytic leukemia–like disease with a lower frequency of leukemia-initiating cells compared with the control. Chromatin immunoprecipitation followed by sequencing revealed a significant reduction in the levels of trimethylation at H3K27 in Ezh2-deficient leukemic cells, not only at Cdkn2a, a known major target of Ezh2, but also at a cohort of genes relevant to the developmental and differentiation processes. Overexpression of Egr1, one of the derepressed genes in Ezh2-deficient leukemic cells, promoted the differentiation of AML cells profoundly. Our findings suggest that Ezh2 inhibits differentiation programs in leukemic stem cells, thereby augmenting their leukemogenic activity.

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Enhanced Autophagy Promotes Survival of Peripheral Blast Cells from Murine Acute Myeloid Leukemia

Blood ◽

10.1182/blood.v124.21.2339.2339 ◽

2014 ◽

Vol 124 (21) ◽

pp. 2339-2339 ◽

Cited By ~ 1

Author(s):

Yoshiki Sumitomo ◽

Junji Koya ◽

Keisuke Kataoka ◽

Takako Tsuruta-Kishino ◽

Ken Morita ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Signaling Pathways ◽

Myeloid Leukemia ◽

Research Funding ◽

Fusion Gene ◽

Leukemic Cells ◽

Cytokine Signaling ◽

Genetic Deletion ◽

Autophagic Activity ◽

Acute Myeloid

Abstract Intensive chemotherapy for the treatment of leukemias inevitably provides cellular and metabolic stress to leukemic cells, leading to programmed cell death. In addition, anti-apoptotic property of leukemic cells could easily induce autophagy in response to anti-leukemic treatments and upregulation of autophagy is likely to contribute to the survival of leukemic cells by driving therapy resistance. Given that bone marrow (BM) supports leukemic cell proliferation by various types of stimuli from stromal cells or leukemic cells themselves, it is supposed that BM-occupying leukemic cells and circulating peripheral leukemic cells would have distinct difference in autophagic activity. However, comprehensive understanding of autophagic activity in leukemic cells has not been achieved so far. Here in this study, autophagic activities of leukemic cells in BM and peripheral blood (PB) from murine acute myeloid leukemia (AML) model driven by MLL-ENL fusion gene were evaluated. In the PB and BM from MLL-ENL AML mice, both mature (CD11b+c-Kit-) and immature (CD11b+c-Kit+) MLL-ENL+ leukemic cells showed no difference in apoptotic status by Annexin-V/DAPI staining regardless of c-Kit expression. By contrast, from cell cycle analysis, c-Kit- leukemic cells in the BM were found to have higher frequency of S/G2 phase than PB counterparts, indicating the proliferative potential of BM leukemic cells. When molecules of intracellular signaling pathways regarding proliferation and survival were assessed, components of MAPK and PI3K-mTOR signaling pathways such as Erk1/2, Akt, S6K, and S6 were highly phosphorylated in c-Kit- BM AML cells compared to c-Kit- PB AML cells, implying the importance of activated cytokine signaling in the BM of MLL-ENL AML. For the autophagic evaluation of PB and BM AML cells, MLL-ENL fusion gene was introduced into autophagy sensor mice, GFP-LC3 transgenic mice, enabling us to check autophagy by GFP. PB AML cells from these mice showed decreased GFP intensity compared to BM counterparts, which meant more activated degradation of autophagosome in peripheral AML cells. By western blotting analysis, PB AML cells had the enhanced conversion of LC3A-I to LC3A-II, indicating activation of autophagy. Furthermore, activation of stress responsive pathways such as nuclear localization of Foxo3a and enhanced phosphorylation of eIF2a was observed in c-Kit- PB AML cells. Interpretation of microarray data using public database (GSE9476 and GSE34577) comparing PB and BM cells from clinical AML samples revealed that PB AML cells have significantly higher expression of amino acid transporters. From these results, it is strongly suggested that peripheral AML cells have enhanced autophagic activity in vivo. To elucidate the functional role of activated autophagy in PB AML cells, MLL-ENL fusion gene was introduced into conditional Atg5 or Atg7 knockout mice (Atg5flox/flox or Atg7flox/flox), both of which are essential for autophagy. Interestingly, genetic deletion of Atg5 or Atg7 in irradiated recipients transplanted with Atg5flox/flox or Atg7flox/flox AML cells caused significant increase of apoptotic cells in peripheral leukemic cells and significant decrease of peripheral white blood cell (WBC) counts and donor cell engraftment in PB, whereas Atg5Δ/Δ or Atg7Δ/Δ MLL-ENL AML mice had neither prolonged survival nor reduced leukemia-initiating capacity in serial transplants. Similar to MLL-ENL AML model, genetic deletion of Atg7 in advanced phase of chronic myeloid leukemia (CML) induced by BCR-ABL plus NUP98-HOXA9 fusion genes caused decreased WBC counts and increased apoptosis in peripheral leukemic cells, indicating that circulating leukemic cells favor autophagy for their survival. It is of note that Atg7 deletion had no impact on apoptosis in normal CD11b+ myeloid fraction, implying that loss of Atg7 could spare normal myeloid cells. It remains to be seen what soluble factors could support the survival of AML cells in BM by suppressing autophagy, but our preclinical data provides the therapeutic potential of autophagy inhibition in the treatment of excessive peripheral leukocytosis. It is also highly expected that inhibition of autophagy could be more effective for BM AML cells when combined with chemotherapeutic agent to activate autophagy. Disclosures Sumitomo: Kyowa Hakko Kirin Co., Ltd.: Employment. Kurokawa:Kyowa Hakko Kirin Co., Ltd.: Research Funding, Speakers Bureau; SHIONOGI & CO., Ltd.: Consultancy; Novartis Pharma K.K.: Consultancy, Research Funding, Speakers Bureau; Celgene K.K.: Consultancy, Speakers Bureau; Bristol-Myers Squibb Company: Consultancy, Research Funding, Speakers Bureau; Sanofi K.K.: Consultancy; CHUGAI PHARMACEUTICAL CO., LTD.: Research Funding, Speakers Bureau; Astellas Pharma Inc., : Research Funding, Speakers Bureau; Dainippon Sumitomo Pharma Co.,Ltd.: Research Funding, Speakers Bureau; Asahi Kasei Co.: Research Funding, Speakers Bureau; Pfizer Inc.: Research Funding; Otsuka Pharmaceutical Co., Ltd.: Research Funding; MSD K.K.: Research Funding, Speakers Bureau; Takeda Pharmaceutical Co.,Ltd.: Research Funding, Speakers Bureau; Nippon Shinyaku Co., Ltd.: Research Funding, Speakers Bureau; TAIHO PHARMACEUTICAL CO., LTD.: Research Funding; Teijin Pharma Limited: Research Funding; Alexion Pharmaceuticals K.K.: Research Funding, Speakers Bureau; Eisai Co., Ltd.: Research Funding, Speakers Bureau; Mitsubishi Tanabe Pharma Corporation: Speakers Bureau; GlaxoSmithKline K.K.: Speakers Bureau; Janssen Pharmaceutical K.K.: Speakers Bureau; Yakult Pharmaceutical Industry Co., Ltd.: Speakers Bureau; Ono Pharmaceutical Co.,Ltd.: Speakers Bureau; Miraca Holdings Inc.: Speakers Bureau; CSL Behring K.K.: Speakers Bureau.

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Immunophenotyping Investigation of Minimal Residual Disease Is a Useful Approach for Predicting Relapse in Acute Myeloid Leukemia Patients

Blood ◽

10.1182/blood.v90.6.2465.2465_2465_2470 ◽

1997 ◽

Vol 90 (6) ◽

pp. 2465-2470 ◽

Cited By ~ 5

Author(s):

J.F. San Miguel ◽

A. Martı́nez ◽

A. Macedo ◽

M.B. Vidriales ◽

C. López-Berges ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Multidrug Resistance ◽

Minimal Residual Disease ◽

Myeloid Leukemia ◽

Residual Disease ◽

Leukemic Cells ◽

Rhodamine 123 ◽

Minimal Residual ◽

Acute Myeloid

A high complete remission rate is currently achieved in patients with acute myeloid leukemia (AML). However, many patients eventually relapse due to the persistence of low numbers of residual leukemic cells that are undetectable by conventional cytomorphologic criteria (minimal residual disease [MRD]). Using immunophenotypic multiparametric flow cytometry, we have investigated in sequential studies (diagnosis and follow-up) the impact of MRD detection on the outcome of 53 AML patients that had achieved morphologic remission with standard AML protocols and displayed at diagnosis an aberrant phenotype. Patients were studied at diagnosis with a panel of 35 monoclonal antibodies in triple staining combinations for detection of aberrant or uncommon phenotypic features. According to these features, a patient's probe was custom-built at diagnosis for the identification of possible residual leukemic cells during follow-up. The level of MRD at the end of induction and intensification therapy correlated with the number of relapses and relapse-free survival (RFS). Thus, patients with more than 5 × 10−3 residual cells (5 residual cells among 1,000 normal bone marrow [BM] cells) identified as leukemic by immunophenotyping in the first remission BM showed a significant higher rate of relapse (67% v 20% for patients with less than 5 × 10−3 residual cells; P = .002) and a lower median RFS (17 months v not reached; P = .01). At the end of intensification, with a cut-off value of 2 × 10−3 leukemic cells, AML patients also separated into two distinct groups with relapse rates of 69% versus 32% (P = .02), respectively, and median RFS of 16 months versus not reached (P = .04). In addition, overall survival was also significantly related to the level of residual cells in the marrow obtained at the end of induction and particularly after intensification therapy (P = .008). Furthermore, we have explored whether residual disease was related with the functional expression of multidrug resistance (MDR-1) at diagnosis as assessed by the rhodamine-123 assay. Patients with ≥5 × 10−3 residual leukemic cells at the end of induction therapy had a significantly higher rhodamine-123 efflux (mean, 56% ± 24%) than those with less than 5 × 10−3 residual cells (mean, 32% ± 31%; P = .04). Finally, multivariate analysis showed that the number of residual cells at the end of induction or intensification therapy was the most important prognostic factor for prediction of RFS. Overall, our results show that immunophenotypical investigation of MRD strongly predicts outcome in patients with AML and that the number of residual leukemic cells correlates with multidrug resistance.

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Post-remission clonal hematopoiesis; Practical implications for measurable residual disease assessment in acute myeloid leukemia (AML).

Journal of Clinical Oncology ◽

10.1200/jco.2020.38.15_suppl.7529 ◽

2020 ◽

Vol 38 (15_suppl) ◽

pp. 7529-7529

Author(s):

Sanam Loghavi ◽

Tomoyuki Tanaka ◽

Ken Furudate ◽

Sa A Wang ◽

Koichi Takahashi

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Residual Disease ◽

Disease Status ◽

Leukemic Cells ◽

Disease Assessment ◽

Individual Gene ◽

Clonal Hematopoiesis ◽

Measurable Residual Disease ◽

Acute Myeloid

7529 Background: Clonal Hematopoiesis may persist following complete remission (CR) in patients with acute myeloid leukemia (AML) but does not necessarily indicate residual AML and may represent persistence of pre-leukemic stem cells. Post-remission CH identified by NGS has not been systemically studied in parallel with measurable residual disease (MRD) detection by flow cytometric immunophenotyping (FCI). Methods: We studied bone marrow sample from AML patients at baseline and CR by targeted deep NGS of 295 genes (median 403x depth) and compared the results to FCI. Measurable residual disease (MRD) detection by FCI was performed by comparing the phenotype at CR to baseline and by detection of leukemia associated immunophenotype (LAIP) and derivation from normal (DFN) (sensitivity: 0.1%). Post-CR CH was defined as presence of mutations originally detected in AML with variant allele frequency > 2.5%. FCI results were categorized into 4 groups: a) AML MRD negative by LAIP or DFN b) AML MRD+ (similar to baseline) c) AML MRD+ (different from baseline), d) Negative for AML MRD, but aberrant phenotype suggestive of pre-leukemic cells. We correlated FCI and NGS results. Results: 101 patients were included in the study. 45 (45%) had persistent post-CR clonal hematopoiesis; 23 (51%) had phenotypic alterations detected by FCI including AML MRD+ in 18 (40%) and pre-leukemic cells in 5 (10%). Among patient with no detectable mutations by NGS (n = 56; 55%), 14 (25%) had FCI aberrancies including AML MRD+ in 4 (7%) and pre-leukemic cells in 10 (18%). CH was significantly more common in samples with residual phenotypic aberrancies detected by FCI (p = 0.004). There was no significant correlation between FCI group d and persistent CH (p = 0.4). Persistent ASXL1 (p = 0.024, OR = 7.2 ) and RUNX1 (p = 0.016; OR = 17.3) mutations were significantly associated with FCI abnormalities. The correlation coefficient between FCI abnormalities and RUNX1 mutations inferred from a Bayesian network structure was 0.66. Conclusions: NGS and FCI are complementary in evaluating post treatment disease status in AML. Post CR-CH is associated with phenotypic abnormalities that either represent residual AML or pre-leukemic cells. The latter may not have the same prognostic implications as AML MRD; however, the association with outcome needs to be elucidated. Single cell DNA sequencing technologies may be helpful in more accurately deciphering the association of individual gene mutations and their contribution to phenotypic aberrations.

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Mitochondrial Mutations as Molecular Markers for Minimal Reidual Disease (MRD) in Acute Myeloid Leukemia.

Blood ◽

10.1182/blood.v104.11.2996.2996 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2996-2996

Author(s):

Alexander A. Morley ◽

Scott A. Grist ◽

Xiao J. Lu ◽

Katrina Patsouris

Keyword(s):

Acute Myeloid Leukemia ◽

Molecular Markers ◽

Myeloid Leukemia ◽

Denaturing Gradient Gel Electrophoresis ◽

Residual Disease ◽

Molecular Techniques ◽

Leukemic Cells ◽

Mitochondrial Mutations ◽

Gradient Gel Electrophoresis ◽

Acute Myeloid

Abstract Measurement of minimal residual disease (MRD) is being increasingly used in haematological malignancies in order to assess prognosis and decide on treatment. However for some patients, including many patients with acute myeloid leukemia (AML), molecular techniques for MRD measurement cannot be performed owing to lack of a molecular marker. We have detected mitochondrial mutations (MM) in the D loop of the mitochondrial genome of the leukemic cells in approximately 40% of patients with AML and have investigated 2 techniques - denaturing gradient gel electrophoresis (DGGE) and single nucleotide primer extension (SNUPE) - to detect and quantify them. Mixing experiments showed that DGGE had a sensitivity of approximately 0.5% for detection of MM, and it was able to detect leukemia in remission marrow in 5 of 6 patients with AML. When present in a patient, MM are usually multiple. We therefore tested a 2-step strategy, as shown in the figure, which involved initial enrichment by PCR using primers directed at 1 or 2 flanking mutations followed by DGGE to detect an inner mutation. In 2 mixing experiments this 2-step strategy increased sensitivity of detection down to 0.0001% (this level of detection was evident in the original gel photo). Figure Figure SNUPE was more sensitive than DGGE and in 3 mixing experiments it showed a sensitivity of 0.02–0.05%. Four patients with AML have now been studied by SNUPE and the levels of MRD in the marrow documenting morphological remission at the end of induction therapy were 2.8%,16.1%,40.8% and 15.7%. The relatively high levels of MRD as measured by PCR in 3 patients suggest that some of the leukaemic cells at the end of induction may be more mature than blasts and thus not identifiable by morphology. While questions remain to be answered, mitochondrial mutations are promising molecular markers for quantifying MRD in AML and possibly other haematological disorders

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Identification of a Set of Seven Genes for the Monitoring of Minimal Residual Disease in Pediatric Acute Myeloid Leukemia.

Blood ◽

10.1182/blood.v106.11.1461.1461 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1461-1461

Author(s):

Daniel Steinbach ◽

Alexander Schramm ◽

Angelika Eggert ◽

Susann Wittig ◽

Nadine Pfaffendorf ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Minimal Residual Disease ◽

Myeloid Leukemia ◽

Residual Disease ◽

Wilms Tumor ◽

Leukemic Cells ◽

Genome Wide ◽

Minimal Residual ◽

Acute Myeloid

Abstract A stepwise approach which combined genome wide expression profiling and a TaqMan realtime PCR based screening was used to identify new markers for the monitoring of minimal residual disease (MRD) in acute myeloid leukemia (AML). Leukemic cells from 52 children with AML were analyzed. Seven genes were identified which are vastly over-expressed in many patients with AML compared to healthy bone marrow: CCL23, GAGED2, MSLN, SPAG6, and ST18 as well as the previously described markers WT1 and PRAME. This set of genes was analyzed in 141 follow-up samples from 25 patients. The expression of all genes decreased to normal levels in patients who achieved a continuous complete remission. Elevated levels of MRD markers were found prior to relapse in 7 out of 10 patients who relapsed. This set of genes should allow a sensitive and specific monitoring of MRD in AML. Notably, some of these markers could also serve as therapeutic targets or might be involved in leukemogenesis. MSLN is already used as a target for immunotherapy in clinical trials in other malignancies. GAGED2 and SPAG6 belong to the family of cancer testis genes which are also studied intensively as targets for immunotherapy. ST18 is a recently discovered tumor suppressor which was not yet described in hematological malignancies. CCL23 is a chemokine that inhibits the proliferation of healthy hematological stem cells. Names, symbols, and geneID of seven MRD markers Gene Symbol Gene Name GeneID CCL23 chemokine (C–C motif) ligand 23 6368 GAGED2 G antigen, family D, 2 9503 MSLN Mesothelin 10232 SPAG6 sperm associated antigen 6 9576 ST18 suppression of tumorigenicity 18 9705 WT1 Wilms tumor 1 7490 PRAME preferentially expressed antigen in melanoma 23532

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Minimal Residual Disease After the First Cycle of Chemotherapy in Acute Myeloid Leukemia: A Comparative Study of WT1 RQ-PCR and 4-Color Flow Cytometry.

Blood ◽

10.1182/blood.v120.21.2535.2535 ◽

2012 ◽

Vol 120 (21) ◽

pp. 2535-2535

Author(s):

Carlo Marani ◽

Raffaella Grasso ◽

Nicoletta Colombo ◽

Marino Clavio ◽

Fabio Guolo ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Residual Disease ◽

Leukemic Cells ◽

Color Flow ◽

Minimal Residual ◽

Acute Myeloid ◽

Intermediate Dose

Abstract Abstract 2535 Background and aims. Detection of minimal residual disease (MRD) has a relevant prognostic value in Acute Myeloid Leukemia (AML). MRD, when used as early treatment response assessment, allows identification of true low-risk and high-risk patients, who may profit alternative chemotherapy approach. In the present retrospective study, we evaluated the impact of MRD assessed by 4-color flow cytometry and WT1 RQ-PCR gene expression in a cohort of AML patients treated at our institution. Methods. Bone marrow samples of 50 adult AML patients (45 de novo and 5 secondary) with available karyotype (K), FLT3-ITD and NPM-A genes mutational status were assessed for MRD after induction. All included patients had a baseline WT1 expression greater than 1000 copies/Ablx104 (range 1060–346060; lab references for normal values 0–500). Fludarabine-based regimen was used as induction; one course of intermediate dose Ara-C 2g/sqm plus idarubicin, followed by 3 courses of intermediate dose Ara-C (2g/sqm) as further consolidation therapy. WT1 log reduction (DWT1) was used to assess the WT1 clearance (DWT1 = logWT1diagnosis – logWT1 post induction). A positive flow MRD was defined by the presence of no less than 25 clustered leukemic cells/105 total events - threshold of 2.5 × 10−4 residual leukemic cells. In patients submitted to bone marrow transplantation (BMT) only the first consolidation course was administered and disease free survival (DFS) was censored at the date of BMT. Results. Two (4%) patients had favorable, 40 (80%) intermediate, and 5 (10%) poor risk K (3 had no metaphases); 14 (28/%) carried FLT3-ITD mutation: among them 8 carried NPM-A mutation too, while 6 were wild type. After the first induction regimen 42 of 50 (84%) patients achieved a complete remission (CR). Patients with a negative flow MRD (32%) had 3 years DFS of 69.5%, whereas those with a positive flow MRD (68%) had a DFS of 27.3% (p = 0.032). Patients with a DWT1 > 1.5 log (65%) had a 3-years DFS of 58.3%, whereas those with a DWT1 ≤ 1.5 log (35%) had a DFS at 1 and 2-years of 13,5% and 0%, respectively (p < 0.001). All patients with a negative flow MRD had also a DWT1 > 1.5 log, whereas 12 (52%) of those who achieved a DWT1 > 1.5 log were still positive by flow MRD. Fourteen (28%) patients with a high risk (HR) profile at diagnosis (poor risk K, intermediate K with FLT3-ITDpos/NPM-Aneg, AML secondary to therapy or previous haematological disorder), 6 were no responder to induction, whereas no one of 8 patients in CR reached a negative MRD status in both test with a very poor outcome (projected DFS 4.8 months). MRD assessment using both flow and DWT1 allow to discriminate no-HR profile patients in three prognostic group: good (flow MRD neg) intermediate (flow MRD pos and DWT1 > 1.5 log) and adverse prognosis (flow MRD pos and DWT1 ≤ 1.5 log) with a projected DFS of 70.5 months, 38.2 months and 4.2 months, respectively (p < 0.001). Conclusions. DWT1 identified patients who would relapse better than flow, whereas a negative flow MRD was the best predictor of long DFS. Using both test in combination with baseline biologic parameters enabled the definition of discrete prognostic categories (Fig 1). Outcome of patients with DWT1 ≤ 1.5 log was very poor and comparable with that of patients with HR profile at diagnosis. In these patients forecast a cure is very difficult with the current treatment option and clinical trials with new drugs should be used already in up-front setting. Disclosures: No relevant conflicts of interest to declare.

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Molecular quantitation of minimal residual disease in acute myeloid leukemia with t(8;21) can identify patients in durable remission and predict clinical relapse

Blood ◽

10.1182/blood.v95.3.815.003k30_815_819 ◽

2000 ◽

Vol 95 (3) ◽

pp. 815-819 ◽

Cited By ~ 102

Author(s):

K. Tobal ◽

J. Newton ◽

M. Macheta ◽

J. Chang ◽

G. Morgenstern ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Residual Disease ◽

Fusion Gene ◽

Clinical Relapse ◽

Rt Pcr ◽

Multicenter Studies ◽

Degradation Rates ◽

Durable Remission ◽

Acute Myeloid

One of the most common translocations in acute myeloid leukemia (AML) is the t(8;21), which produces the fusion gene AML1-MTG8. We have developed a sensitive competitive reverse transcriptase-polymerase chain reaction (RT-PCR) assay forAML1-MTG8 transcripts, coupled with a competitive RT-PCR for the ABL transcript as a control to accurately estimate the level of amplifiable RNA. We have shown that AML1-MTG8 andABL transcripts have equal degradation rates. Thus, this method is useful for multicenter studies. We studied 25 patients with t(8;21) AML by means of serial analysis done on bone marrow (BM) and peripheral blood (PB) samples from 21 patients. Our analysis showed that, in general, a successful induction chemotherapy produces a reduction of 2 to 3 log in the level of AML1-MTG8, followed by a further 2 to 3 log after consolidation/intensification chemotherapy. Levels up to 1 × 103 and 1 × 102 molecules/μg of RNA in BM and PB, respectively, were compatible with durable remission. On the other hand, 5 patients with levels of 0.71 × 105to 2.27 × 105 molecules/μg of RNA in BM and 2.27 × 103 to 2.27 × 104 molecules/μg of RNA in PB had hematologic relapse within 3 to 6 months. Our data indicate that serial quantitation of AML1-MTG8 transcripts is useful in identifying patients at high risk of relapse and may offer an opportunity for clinical intervention to prevent hematologic relapse. This approach was applied successfully in a patient who had an allogeneic BM transplantation. We also suggest that PB may be used an alternative to BM for quantitating AML1-MTG8 transcripts.

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