March 2023 – Hematopathology & Laboratory Genetics and Genomics

These results support the presence of a cytogenetically cryptic and atypical MLLT10::KMT2A fusion which is associated with an unfavorable prognosis in pediatric AML.

Pediatric acute myeloid leukemia (AML) is a rare and genetically heterogeneous disorder. AML occurring in infants less than 1-2 years of age constitutes a specific subgroup of disease. Amongst others, this entity is characterized by a distinct genetic profile with rearrangements involving the KMT2A (formerly known as MLL) gene at 11q23 representing the most frequent cytogenetic alteration in this patient population. These are documented in approximately 50% of cases.

KMT2A rearrangements juxtapose KMT2A, a gene involved in transcriptional regulation and chromatin modification, with one of numerous partner genes, yielding a chimeric fusion oncoprotein which contributes to oncogenesis via several pathways such as enhanced transcriptional activity and aberrant promoter-like epigenetic signatures. Common KMT2A rearrangement partners in pediatric AML include, amongst others, MLLT3, MLLT10, MLLT4, ELL,MLLT1, AFF1, and MLLT11.

Classically, KMT2A rearrangements occur through reciprocal translocation events. These structural abnormalities may be detectable by conventional chromosome analysis depending on the size of the translocated segments and the achieved resolution of the chromosome banded study. On FISH analysis, KMT2A rearrangements can be detected with either 1) break-apart probes (BAPs), where a disruption of KMT2A is seen as single separate red (R) and green (G) signals, or 2) dual-color dual-fusion probes, where specific KMT2A fusions are seen by FISH patterns showing 1 red (R), 1 green (G), and 2 fusion (F) signals, which are typical findings of reciprocal KMT2A translocations. However, more complex or atypical rearrangements involving KMT2A are also observed. These include, amongst others, subtle insertions of KMT2A segments into other chromosomes and of partner gene segments into KMT2A, inversion events, complex translocations, or translocations with accompanying deletions. Thus, these atypical rearrangements may elude detection by some conventional cytogenetics methodologies (i.e., be “cytogenetically cryptic”) and show atypical FISH probe signal patterns, as seen in this case.

The presented case serves as an example of such cryptic KMT2A rearrangements as no abnormalities were apparent on conventional chromosome analysis and on KMT2A BAP FISH. Nonetheless, a KMT2A::MLLT10 fusion signal (with a 1R2G1F-pattern) was detected with dual-color dual-fusion FISH, a finding which is in support of a KMT2A rearrangement. Compared to KMT2A BAP, dual-color dual-fusion probe strategies are more sensitive and specific, as they label KMT2A and its target rearrangement partner in different colors. While the mechanisms underlying the observed  pattern in the presented case cannot be ascertained in the absence of higher-resolution genetic testing, it could be attributable to a small-size insertion of MLLT10 into KMT2A. Small insertions of partner regions within KMT2A preclude sufficient separation of the KMT2A BAP FISH probes to be detected, thus remaining below the resolution of conventional chromosome analysis and BAP FISH probes.

Accordingly, the results obtained in this case support the presence of a MLLT10::KMT2A fusion event, which was cryptic by G-banding analysis and with KMT2A BAP FISH. While KMT2A rearrangements with different rearrangement partners may variably impact prognosis, KMT2A::MLLT10 rearrangements are generally considered to confer unfavorable prognostic implications in pediatric AML.

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