Coexistence of ETV6-RUNX1 and MLL aberration among pediatric acute lymphoblastic leukemia: case reports and a literature review
Highlight box
Key findings
• Cases with an MLL-partial tandem duplications (MLL-PTD) or other types of MLL aberration are in fact a distinct subentry among ETV6-RUNX1 B-cell acute lymphoblastic leukemia (B-ALL).
What is known and what is new?
• It is known that ETV6-RUNX1 is usually related to favorable prognosis, but MLL aberration has demonstrated poor prognosis among pediatric ALL.
• Herein, we reported 4 cases of the coexistence of ETV6-RUNX1 and MLL-PTD in pediatric ALL patients and showed the favorable outcome.
What is the implication, and what should change now?
• This result also indicates a simple explanation for the good clinical characteristics and good prognosis about coexistence of ETV6-RUNX1 and MLL aberration.
Introduction
In pediatric acute lymphoblastic leukemia (ALL), ETV6-RUNX1 is the most common genetic abnormality, which is found in 25% of pediatric ALL cases, and it is usually related to favorable prognosis (1-3). MLL aberrations have been described in a small number of ALL cases and are considered a poor prognostic marker (4-6). However, the coexistence of ETV6-RUNX1 and MLL aberrations in pediatric ALL patients is rare. Herein, we report 4 cases of pediatric ALL with the coexistence of ETV6-RUNX1 and MLL aberration who were diagnosed from February 2008 to June 2016. We present this article in accordance with the CARE reporting checklist (available at https://tcr.amegroups.com/article/view/10.21037/tcr-23-142/rc).
Case presentation
Case reports
All procedures performed in this study were in accordance with the ethical standards of the institutional and/or national research committees and with the Helsinki Declaration (as revised in 2013). Written informed consent was provided by the patients’ parents/legal guardians for publication of these case reports. A copy of the written consent is available for review by the editorial office of this journal. The study protocol and informed consent were approved by the Ethics Committee of Sun Yat-Sen Memorial Hospital (No. SYSKY-2022-333-01).
Case 1
An 8-year-old male presented with a week of pallor and coughing on February 28, 2012. Blood routine test showed white blood cell (WBC) count 5×109/L, hemoglobin (Hb) 104 g/L, and platelet (PLT) 28×109/L. Flow cytometry (FC) on bone marrow (BM) revealed findings consistent with a B-cell ALL (B-ALL) and CD10 was positive. A normal male karyotype was shown by cytogenetic analysis, and fluorescence in situ hybridization (FISH) studies showed that (12;21) (p13;q22)/ETV6-RUNX1 in 92% of the interphase nuclei and MLL aberration was negative. However, reverse transcription-polymerase chain reaction (RT-PCR) showed an MLL exon 6–2 fusion. The case was included in the Guangdong-2008-ALL protocol for induction chemotherapy. He exhibited prednisone good response (PGR) on day 8, and FC performed on day 15 BM showed minimal residual disease (MRD) B-ALL (4% lymphoblasts) and on day 33 was negative for residual disease. Later, the patient was unable to continue treatment because a lack of family financial resources, and treatment was abandoned 1 month after diagnosis.
Case 2
In case 2, another 8-year-old male childhood patient was admitted to hospital for a month of pallor and a 5-day of fever on April 14, 2011. The results of routine blood examination revealed a WBC of 22.58×109/L, Hb of 50 g/L, and PLT of 22×109/L. BM indicated that primitive lymphoid and immature lymphocytes accounted for 98.5%. The result of BM FC was consistent with that of B-ALL and CD10 was positive. We used an ETV6-RUNX1 probe and an MLL probe with FISH analysis on metaphase and interphase cells revealed ETV6-RUNX1 (82% of cells), whereas FISH analysis revealed negative MLL aberration. Nevertheless, RT-PCR showed an MLL exon 5–2 fusion. He exhibited PGR on day 8, and FC performed on day 15 and on day 33 BM were negative for MRD. The patient was also treated with Guangdong-2008-ALL protocol and achieved complete response (CR), which, at the time of writing, had been maintained for 10 years.
Case 3
This child was a 3-year-old male who developed fever for 6 days and abnormal hemogram for 2 days on March 23, 2012. Blood routine examination showed Hb 62 g/L, PLT 32×109/L, and WBC 4.67×109/L, but 96.5% of the primordial cells were of different sizes. BM FC confirmed that he had B-ALL and CD10 was positive. ETV6-RUNX1 were positive with 86% by FISH analysis, and no MLL was found, but RT-PCR confirmed exon 5–2 of MLL was fused. The patient received induction chemotherapy with Guangdong-2008-ALL regimen. He exhibited PGR on day 8, BM FC showed little B-ALL residue on day 15 (6% of lymphoblasts), and MRD was negative on day 33. Consolidation chemotherapy was well tolerated and there was no acute reaction. He was in remission during a 9-year follow-up and is still in remission.
Case 4
Case 4, a 5-year-old male patient hospitalized due to swelling and pain in the right wrist for 2 weeks on October 21, 2010. Routine blood examinations showed that WBC 3.5×109/L, Hb 88 g/L, PLT 124×109/L. BM cytology indicated that blast cells accounted for 98%. Cytogenetic analysis showed a normal male karyotype; FISH analysis showed ETV6-RUNX1 was positive, MLL aberration was negative, and CD10 was positive. Nonetheless, RT-PCR showed an MLL exon 5, 6–2 fusion. The patient was treated with Guangdong-2008-ALL protocol for induction chemotherapy. He exhibited PGR on day 8, and BM FC performed on day 15 showed MRD (4% lymphoblasts) and on day 33 was negative for MRD. To date, he has been in CR for 11 years since diagnosis. Table 1 shows the patients’ characteristics.
Table 1
| Case | Age (years) | Gender | WBC | Prednisone response | BM blasts on day 15 | BM blasts on day 33 | Fusion gene | Outcome (time since diagnosis) | |
|---|---|---|---|---|---|---|---|---|---|
| ETV6/RUNX1 | MLL | ||||||||
| 1 | 8.3 | M | 5×109/L | PGR | M1 | M1 | 92% (+) | MLL-PTD (ex6/MLLex2) | Abandon treatment |
| 2 | 8.2 | M | 22.58×109/L | PGR | M1 | M1 | 82% (+) | MLL-PTD (ex5/MLLex2) | CR for 10 years |
| 3 | 3.3 | M | 4.67×109/L | PGR | M2 | M1 | 86% (+) | MLL-PTD (ex5/MLLex2) | CR for 9 years |
| 4 | 5.2 | M | 3.5×109/L | PGR | M1 | M1 | 80% (+) | MLL-PTD (ex5,6/MLLex2) | CR for 11 years |
M1: bone marrow blast <5%; M2: bone marrow blast of 5–25%. “+” means positive. ALL, acute lymphoblastic leukemia; WBC, white blood cell; BM, bone marrow; M, male; PGR, prednisone good response; PTD, partial tandem duplications; CR, complete remission.
Literature review
Using the keywords “ETV6-RUNX1”, “MLL”, “children” and “acute lymphoblastic leukemia”, a literature search of coexistence of ETV6-RUNX1 and MLL aberration was performed in the database of PubMed, and 4 articles were retrieved finally [Attarbaschi et al. (7), 2007; Amare Kadam et al. (8), 2008; Matthew C. Hiemenz et al. (9), 2011; Gagnon et al. (10)]. Among the 4 reports, the patient age range was from 2 to 7 years, including 9 males and 7 females and the WBC count was (2.66–68.6)×109/L. When it comes to fusion genes, they all had ETV6/RUNX1-positive (39–94% of cells). Among them, 8 ETV6/RUNX1-positive cases exhibited MLL deletion, and 2 cases had der(21) duplication.
Attarbaschi et al. (7) reported that 7 cases of concurrent ETV6-RUNX1 and MLL aberration (case 1 to case 7 in Table 2) among a cohort of 824 B-ALL between 22 September 1986 and 31 May 2005 in 4 center trials of Berlin-Frankfurt-Münster (BFM) group, and the frequency of coexistence of ETV6-RUNX1 and MLL aberration was 0.8% (7/824). The 8-year RFS and OS were 86%±13% and 100% for the 7 cases, respectively.
Table 2
| Case | Age (years) | Gender | WBC | Prednisone response | BM blasts on day +15 | BM blasts on day +33 | Fusion gene | Outcome (time since diagnosis) | |
|---|---|---|---|---|---|---|---|---|---|
| ETV6/RUNX1 | MLL | ||||||||
| 1 | 5 | F | 25×109/L | PGR | M2 | M1 | 83% (+) | MLL deletion | Relapse in ovary (4.13 yr) |
| 2 | 2 | F | 29×109/L | PGR | M1 | M1 | 75% (+) | MLL deletion | Secondary AML (4.5 yr) |
| 3 | 4 | F | 15×109/L | PGR | M1 | M1 | 83% (+) | MLL deletion | CR (8 yr) |
| 4 | 7 | F | 7×109/L | PGR | M2 | M1 | 80% (+) | MLL deletion | CR (8 yr) |
| 5 | 3 | M | 5×109/L | PGR | M2 | M1 | 75% (+) | MLL deletion | CR (8 yr) |
| 6 | 5 | M | 10.1×109/L | PGR | M2 | M1 | 92% (+) | MLL deletion | CR (8 yr) |
| 7 | 2 | M | 13.5×109/L | PGR | M1 | M1 | 86% (+) | MLL deletion | CR (8 yr) |
| 8 | 5 | M | 13.3×109/L | PGR | M1 | M1 | 75% (+) | MLL deletion | CR (1 yr 3 mo) |
| 9 | 4 | M | 17.9×109/L | PGR | M1 | M1 | 39% (+) | Southern-positive MLL aberration | CR (6 yr 8 mo) |
| 10 | 7 | M | 2.6×109/L | PGR | M2 | M1 | 79% (+) | 27% monoallelic loss | CR (10 mo) |
| 11 | 2 | M | 8×109/L | PGR | M1 | M1 | 72% (+) | 41% monoallelic loss | Induction death |
| 12 | 3 | M | 2.5×109/L | PGR | M1 | M1 | 70% (+) | 26% monoallelic loss | CR (2 yr 4 mo) |
| 13 | 4 | F | 4.9×109/L | PGR | M1 | M1 | 90% (+) | 66% monoallelic loss | CR (3 yr 5 mo) |
| 14 | 4 | F | 68.6×109/L | PGR | M1 | M1 | 94% (+) | 15% monoallelic loss | CR (2 yr 5 mo) |
| 15 | 3 | M | 10.7×109/L | PGR | M1 | M1 | 58% (+) | Southern-positive MLL aberration | CR (6 mo) |
| 16 | 3 | F | 10.95×109/L | PGR | M1 | M1 | 42% (+) | AF9 | CR (3 mo) |
ALL, acute lymphoblastic leukemia; WBC, white blood cell; BM, bone marrow; F, female; M, male; PGR, prednisone good response; yr, years; mo, months; AML, acute myeloid leukemia; CR, complete remission.
Amare Kadam et al. (8) reported 7 cases (case 8 to case 14 in Table 2) among 428 childhood B-cell precursor ALL (BCP-ALL) patients (including 76 ETV6/RUNX1-positive cases) who were treated at their center from 2000 to 2006, and the frequency of coexistence of ETV6-RUNX1 and MLL aberration was 1.6% (7/426). The 3-year event-free survival (EFS) and overall survival (OS) were both 85.7%.
Hiemenz et al. (9) described a 3-year-old male who presented with 3 days of fever, pallor, and lymphadenopathy. Laboratory examination revealed Hb of 77 g/L, PLT of 60×109/L, and normal WBC of 10.7×109/L, but with 41% blasts of variable cell size. FC on BM revealed findings consistent with a B-ALL. Cytogenetic analysis showed a normal male karyotype; FISH studies showed (12;21)(p13;q22)/ETV6-RUNX1 in 58% of the interphase nuclei and a small clonal population (3% of the cells) with an MLL rearrangement.
Gagnon et al. (10) described a 3-year-old female patient with ALL for whom complete blood count revealed significant cytopenia with a Hb level of 18 g/L and total WBC count of 10.95×109/L. FISH studies using a dual color fusion probe for MLL-AF9 were performed and revealed MLL-AF9 fusion signals in 40% of interphase nuclei and in 2 metaphase cells. Additional abnormalities were also observed including an ETV6-RUNX1 fusion in 42% of nuclei.
Discussion
In general, ETV6-RUNX1 and MLL aberration are considered disease-initiating primary genetic lesions in B-ALL with prevailing mutual exclusivity. However, we found that the frequency of coexistence of the 2 fusion genes in our hospital was 0.98%, which was consistent with Attarbaschi et al.’s report (7) (0.8%), but was lower than reported by Amare Kadam et al. (8) (1.6%). Our exploration of the reason revealed that it was not only due to different sample sizes, but also the respective screening approaches. The detection of MLL cases relies on the diagnostic procedures used in studies, because traditional cytogenetics or MLL FISH technology (which will show typical MLL translocations) would confirm them, whereas examining ETV6-RUNX1 with RT-PCR or FISH alone would miss those (11). Our study showed that children who showed co-expression of fusion gene were MLL aberration-negative for FISH, but MLL-partial tandem duplications (PTD) was detected by RT-PCR.
The MLL dichromatic break aberration/separation aberration probe used in this study can only detect all translocation aberrations of 11q23 MLL gene, and cannot be used for detection of other aberration types such as deletion, inversion, and repetition. Conversely, RT-PCR can quickly and accurately detect multiple fusion genes, detect the fusion gene type formed by chromosome translocation in MLL gene aberration, PTD, and detect the coexistence of many fusion genes at the same time, which is a feasible method to detect MLL gene aberration (12). Thereby, we suggest that the joint detection of fusion genes by the 3 methods should be recommended.
Considering the coexistence of 2 recurrent genetic abnormalities, the clinical course of these cases is expected to be poor because the clinical outcome of ALL patients with MLL aberration is poor (13-15). However, combining the cases in our study and those reported in the literature, the 20 patients revealed favorable clinical and laboratory features at diagnosis. The association between these patients is that they were all between age 2 to 8 years, with low WBC and PGR on day 8, day 15, and day 33, with a similar favorable EFS and OS rate. This result of coexistence of the 2 fusion genes seemed to be related to ETV6-RUNX1 rather than MLL aberration, which indicated that ETV6-RUNX1 was dominant, whereas MLL aberration was inhibited. Based on the current data, we suggest that the interaction and expression of ETV6-RUNX1 protein with various cellular signaling pathways may affect pharmacological reaction, but have nothing to do with any minor events of MLL. The interaction between ETV6-RUNX1 protein products and abnormal MLL and its effect on cell environment, and if MLL plays a cancer inhibitory role due to allele deletion in ETV6-RUNX1 positive environment warrants further study (16). Another possible explanation for the good outcome of coexistence of ETV6-RUNX1 and MLL aberration is that MLL aberration merely represents a highly specific nonrandom secondary genetic abnormality that should always provide an alert to the potential presence of an ETV6/RUNX1-associated pathology that is rarely identified with conventional cytogenetic means only. In contrast to the previous study, Gagnon et al. (10) used whole genome sequencing to elucidate and characterize this case of coexistence, and they revealed that the apparent fusion between MLL and AF9 by FISH was not predicted to result in a bona fide MLL-AF9 fusion and would not be expected to alter the intrinsic MLL regulatory mechanisms and confer leukemogenic potential, whereas it would be expected to result in a lack of protein production from this allele.
The biggest difference between our study and the previous literature was that in the previous literature, MLL aberration involved MLL deletion, southern-positive MLL aberration, and monoallelic loss, but our study showed that 4 ETV6/RUNX1-positive cases had MLL-PTD. MLL-PTD is one of the common forms of MLL gene aberration, which is the product of partial insertion of the 5' end of the MLL gene into the genome and self-fusion (17). It has been reported that MLL-PTD positive patients easily relapse after the first CR, the survival time is short, and the prognosis is very poor (18,19). MLL-PTD can be detected in peripheral blood, BM, and umbilical cord blood of healthy individuals, which indicates that MLL-PLD is not a decisive factor in the occurrence of MLL-PTD-related leukemia, and may provide a genetic background for subsequent changes in key genes of ALL (20). In our study, 3 patients who exhibited coexistence of ETV6-RUNX1 and MLL aberration achieved CR and had a long-term EFS, which is inconsistent with the previous studies (7-10).
Conclusions
We believe that the results of our analysis primarily provide compelling evidence that cases with an MLL-PTD or other types of MLL aberration comprise a distinct subentity among ETV6-RUNX1 B-ALL. Therefore, this finding also indicates a simple explanation for their good clinical characteristics and good prognosis.
Acknowledgments
Funding: This work was supported by
Footnote
Reporting Checklist: The authors have completed the CARE reporting checklist. Available at https://tcr.amegroups.com/article/view/10.21037/tcr-23-142/rc
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Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tcr.amegroups.com/article/view/10.21037/tcr-23-142/coif). The authors have no conflicts of interest to declare.
Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All procedures performed in this study were in accordance with the ethical standards of the institutional and/or national research committees and with the Helsinki Declaration (as revised in 2013). Written informed consent was provided by the patients’ parents/legal guardians for publication of these case reports. A copy of the written consent is available for review by the editorial office of this journal. The study protocol and informed consent form were approved by the Ethics Committee of Sun Yat-Sen Memorial Hospital (No. SYSKY-2022-333-01).
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References
- Bang B, Eisfeldt J, Barbany G, et al. A somatic UBA2 variant preceded ETV6-RUNX1 in the concordant BCP-ALL of monozygotic twins. Blood Adv 2022;6:2275-89. [Crossref] [PubMed]
- Sun C, Chang L, Liu C, et al. The study of METTL3 and METTL14 expressions in childhood ETV6/RUNX1-positive acute lymphoblastic leukemia. Mol Genet Genomic Med 2019;7:e00933. [Crossref] [PubMed]
- Qiu KY, Xu HG, Luo XQ, et al. Prognostic Value and Outcome for ETV6/RUNX1-Positive Pediatric Acute Lymphoblastic Leukemia: A Report From the South China Children's Leukemia Group. Front Oncol 2021;11:797194. [Crossref] [PubMed]
- Qiu KY, Zhou DH, Liao XY, et al. Prognostic value and outcome for acute lymphocytic leukemia in children with MLL rearrangement: a case-control study. BMC Cancer 2022;22:1257. [Crossref] [PubMed]
- Britten O, Ragusa D, Tosi S, et al. MLL-Rearranged Acute Leukemia with t(4;11)(q21;q23)-Current Treatment Options. Is There a Role for CAR-T Cell Therapy? Cells 2019;8:1341. [Crossref] [PubMed]
- Molugu K, Battistini GA, Heaster TM, et al. Label-Free Imaging to Track Reprogramming of Human Somatic Cells. GEN Biotechnol 2022;1:176-91. [Crossref] [PubMed]
- Attarbaschi A, Mann G, Strehl S, et al. Deletion of 11q23 is a highly specific nonrandom secondary genetic abnormality of ETV6/RUNX1-rearranged childhood acute lymphoblastic leukemia. Leukemia 2007;21:584-6. Erratum in: Leukemia 2007;21:1135. [Crossref] [PubMed]
- Amare Kadam PS, Raje GC, Pais AP, et al. Coexistence of ETV6/RUNX1 and MLL aberrations in B-cell precursor acute lymphoblastic leukemia discloses a small subclass of BCP-ALL. Cancer Genet Cytogenet 2008;182:27-32. [Crossref] [PubMed]
- Hiemenz MC, Chen W, Winick N, et al. Coexistence of t(12;21)(p13;q22)/ETV6-RUNX1 and 11q23/MLL Rearrangement in B Acute Lymphoblastic Leukemia: A Case Report and Review of the Literature. J Assoc Genet Technol 2011;37:213-5. [PubMed]
- Gagnon MF, Smadbeck JB, Sharma N, et al. Apparent coexistence of ETV6::RUNX1 and KMT2A::MLLT3 fusions due to a nonproductive KMT2A rearrangement in B-ALL. Leuk Lymphoma 2022;63:2243-6. [Crossref] [PubMed]
- Mollstedt J, Mansouri L, Rosenquist R. Precision diagnostics in chronic lymphocytic leukemia: Past, present and future. Front Oncol 2023;13:1146486. [Crossref] [PubMed]
- Zou P, Zhou M, Wen J, et al. The long-term outcome and risk factors for precursor B cell acute lymphoblastic leukemia without specific fusion genes in Chinese children: experiences from multiple centers. Bosn J Basic Med Sci 2022;22:238-46. [PubMed]
- Spatari G, Allegra A, Carrieri M, et al. Epigenetic Effects of Benzene in Hematologic Neoplasms: The Altered Gene Expression. Cancers (Basel) 2021;13:2392. [Crossref] [PubMed]
- Fang Y, Yang C, Teng D, et al. Discovery of higenamine as a potent, selective and cellular active natural LSD1 inhibitor for MLL-rearranged leukemia therapy. Bioorg Chem 2021;109:104723. [Crossref] [PubMed]
- Kundu A, Kowarz E, Marschalek R . The role of reciprocal fusions in MLL-r acute leukemia: studying t(6;11) fusion proteins. Klinische Pediatrie 2020. doi:
10.1055/s-0040-1709763 .10.1055/s-0040-1709763 - Li M, Jones L, Gaillard C, et al. Initially disadvantaged, TEL-AML1 cells expand and initiate leukemia in response to irradiation and cooperating mutations. Leukemia 2013;27:1570-3. [Crossref] [PubMed]
- Bäsecke J, Podleschny M, Clemens R, et al. Lifelong persistence of AML associated MLL partial tandem duplications (MLL-PTD) in healthy adults. Leuk Res 2006;30:1091-6. [Crossref] [PubMed]
- Bernot KM, Siebenaler RF, Whitman SP, et al. Toward personalized therapy in AML: in vivo benefit of targeting aberrant epigenetics in MLL-PTD-associated AML. Leukemia 2013;27:2379-82. [Crossref] [PubMed]
- Sun QY, Ding LW, Tan KT, et al. Ordering of mutations in acute myeloid leukemia with partial tandem duplication of MLL (MLL-PTD). Leukemia 2017;31:1-10. [Crossref] [PubMed]
- Nasilowska-Adamska B, Solarska I, Paluszewska M, et al. FLT3-ITD and MLL-PTD influence the expression of MDR-1, MRP-1, and BCRP mRNA but not LRP mRNA assessed with RQ-PCR method in adult acute myeloid leukemia. Ann Hematol 2014;93:577-93. [Crossref] [PubMed]
