Unveiling a Novel Fusion circRNA Regulating Mitochondrial Metabolism in t(4;11) B-cell Acute Lymphoblastic Leukemia

Highlight

The t(4;11) translocation produces a fusion gene KMT2A::AFF1 associated with poor prognosis in B-cell Acute Lymphoblastic Leukemia (B-ALL). Researchers have identified a novel fusion circular RNA (circRNA), termed AK_7_3, derived from the KMT2A::AFF1 gene, which plays an anti-apoptotic role by modulating mitochondrial function in leukemic cells. This circRNA is recurrently detected in pediatric and adult t(4;11)-positive B-ALL patients and impacts oxidative stress response, apoptosis regulation, and mitochondrial bioenergetics.

Study Background

B-cell Acute Lymphoblastic Leukemia (B-ALL) is a hematologic malignancy characterized by the malignant proliferation of immature B-cell precursors. Among genetic abnormalities defining B-ALL subtypes, the t(4;11)(q24;q23) translocation is a hallmark of a high-risk subset found both in pediatric and adult patients. This translocation creates the KMT2A::AFF1 fusion gene, which is implicated in leukemogenesis and poor clinical outcomes. Despite progress in characterizing linear fusion transcripts, the role of circular RNAs (circRNAs)—a class of stable, covalently closed RNA molecules formed by back-splicing—in the pathobiology of t(4;11) B-ALL has remained unclear. Fusion circRNAs (f-circRNAs) may exert regulatory functions influencing the survival and metabolism of leukemic cells, but detailed mechanistic insights are limited.

Study Design

This study employed molecular and cellular biology approaches to identify and characterize a novel fusion circRNA generated from the KMT2A::AFF1 fusion gene. The authors examined three human B-ALL cell lines (SEM, RS4;11, ALL-PO), all harboring the t(4;11) translocation, and clinical samples from pediatric and adult t(4;11)-positive B-ALL patients at diagnosis and relapse. The novel back-splicing junction joining AFF1 exon 7 to KMT2A exon 3 (AK_7_3) was detected by reverse transcription PCR and validated by sequencing. Functional studies included knockdown of AK_7_3 in vitro, with downstream transcriptome analyses by RNA-Seq, assessment of apoptosis, mitochondrial morphology and function measured by confocal microscopy, Seahorse bioenergetics assays, and electron microscopy.

Key Findings

The study discovered a novel, recurrent fusion circRNA, AK_7_3, generated by the back-splicing of AFF1 exon 7 to KMT2A exon 3 in t(4;11)-positive B-ALL cell lines and patient samples. AK_7_3 was present in 12/18 pediatric and 17/24 adult cases, including paired diagnosis and relapse specimens, but absent in t(4;11)-negative controls, indicating its high specificity and recurrence. The fusion circRNA exists as two splicing variants linked to exon 4 of KMT2A, also seen in the linear fusion transcript.

Functional knockdown of AK_7_3 in SEM cells produced a significant upregulation of genes involved in oxidative stress responses and apoptosis regulation, underscoring AK_7_3’s role as an anti-apoptotic factor. Consistent with transcriptomic data, experimental silencing of AK_7_3 increased leukemic cell apoptosis, demonstrating its survival-promoting function.

At the mitochondrial level, AK_7_3 knockdown induced an overproduction of reactive oxygen species (ROS), elevated mitochondrial membrane potential, and significant enhancement of mitochondrial respiration, along with ultrastructural abnormalities observed via electron microscopy. These findings suggest that AK_7_3 modulates mitochondrial metabolism and integrity to support leukemic cell survival.

Expert Commentary

This study advances our understanding of the molecular biology underpinning t(4;11)-driven B-ALL by unveiling the novel fusion circRNA AK_7_3 with critical functional effects on mitochondrial metabolism and apoptosis resistance. Fusion circRNAs represent an emerging class of regulatory RNA molecules with oncogenic potential. The demonstration that AK_7_3 contributes to mitochondrial homeostasis provides a mechanistic link between fusion gene expression and metabolic adaptation in leukemia.

Given the poor prognosis of t(4;11) B-ALL and resistance to conventional chemotherapy, AK_7_3 may serve as both a biomarker and a potential therapeutic target. Targeting circRNAs is still nascent, but approaches such as antisense oligonucleotides or RNA interference could be explored to disrupt the anti-apoptotic and metabolic advantages conferred by this fusion circRNA. However, further in vivo validation and assessment of potential off-target effects are warranted.

The study is limited primarily by its in vitro and ex vivo design, and clinical correlations with response and survival outcomes remain to be elucidated. Exploration of AK_7_3 expression dynamics during therapy and relapse might reveal insights into clonal evolution and resistance mechanisms.

Conclusion

The identification of the KMT2A::AFF1-derived fusion circRNA AK_7_3 enriches understanding of t(4;11) B-ALL biology by linking a novel circRNA to mitochondrial function and apoptotic regulation. The circRNA promotes leukemic cell survival by maintaining mitochondrial integrity and reducing oxidative stress-induced apoptosis, which likely contributes to disease aggressiveness. This study highlights the potential of fusion circRNAs as novel molecular players in leukemia pathogenesis and as future therapeutic targets to improve outcomes in high-risk B-ALL.

Funding and ClinicalTrials.gov

The original study did not specify funding details or clinical trial registration within the abstract. Further information could be accessed through the full publication in Haematologica.

References

1. Tolomeo D, Bardini M, Venuto S, et al. A novel KMT2A::AFF1-derived fusion circRNA regulates mitochondrial metabolism in t(4;11) B-cell acute lymphoblastic leukemia. Haematologica. 2026 Aug 13. PMID: 42610417.
2. Meyer C, Burmeister T, Gröger D, et al. The MLL recombinome of acute leukemias in 2017. Leukemia. 2018 Apr;32(5):273-284.
3. Salzman J. Circular RNA Expression: Its Potential Regulation and Function. Trends Genet. 2016 May;32(5):309-16.
4. Lasda E, Parker R. Circular RNAs: diversity of form and function. RNA. 2014 Dec;20(12):1829-42.
5. Fonseca PM, Gruber Mazzucchelli CA, et al. Mitochondrial metabolism and leukemogenesis: potential treatments targeting energy metabolism and bioenergetics in acute leukemia. Blood Rev. 2020 Sep;43:100669.