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Institute of Clinical Pathology and Medical Research, New South Wales Health Pathology, Westmead Hospital, Westmead, NSW, AustraliaUniversity of Sydney, Westmead Medical School, Westmead, NSW, Australia
However, KMT2A amplification is rarer and generally reported to occur in only 1% of cases of acute myeloid leukaemia (AML). KMT2A amplification is rarely reported in B-lymphoblastic leukaemia (B-ALL) with only a handful of cases
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
in the literature. To our knowledge, we present only the second case report of KMT2A amplification with corresponding TP53 deletion in adult de novo CD10 negative B-ALL.
A 69-year-old male presented to the emergency department with symptoms of melena, coffee ground vomit, syncope and confusion, on a background of 2 weeks of abdominal pain. His only other significant medical history was consumption of 8–10 beers per day. He was sent for an urgent gastroscope which showed sigmoid diverticulitis with localised perforation. He denied any past significant medical history. His FBC results showed he was pancytopenic (Hb 52 g/L, WCC 3.9×109/L, Neut 0.5×109/L and platelet count 7×109/L) with 17% circulating blasts (Fig. 1). Flow cytometry was performed on the peripheral blood and showed that the blasts were positive for CD45(dim), CD19, CD34, CD38, CD15(variable) and HLA-DR. They were negative for CD3, CD4, CD5, CD7, CD10, CD13, CD20,CD33,CD56,CD64,CD117.
Fig. 1Blasts consist of large cells with a high N:C ratio, variable nuclear morphology (some with folded nuclei), intermediate chromatin, multiple nucleoli, and a scant quantity of basophilic cytoplasm. No cytoplasmic granules or Auer rods are seen.
Plans were made for a bone marrow biopsy the following day, however the patient self-discharged from hospital, accepted referral to palliative care and died shortly after.
As a result of this and the prospect of no bone marrow sample, additional flow cytometry testing was performed on the peripheral blood. The additional flow cytometry markers showed the blasts to be positive for intracellular CD79a, CD22 and negative for MPO and intracellular CD3, confirming B cell lineage. At the same time rapid fluorescence in situ hybridisation (FISH)
for BCR::ABL1 gene fusion and a KMT2A rearrangement was performed on directly harvested peripheral blood cells using the LSI BCR/ABL dual colour dual fusion (DCDF) probe and the LSI MLL (KMT2A) dual colour break-apart (DCBA) probe (Abbott Molecular, USA) to further classify the leukaemia/allow access to targeted therapy depending upon the results.
The FISH showed KMT2A amplification with >20 signals per cell (Fig. 2A–C). There was no evidence of BCR::ABL1 gene fusion; however, there were three to four copies of BCR and three copies of ABL1 suggesting aneuploidy for chromosomes 9 and 22 (Fig. 2D). Once the KMT2A amplification result was known, a diagnosis of AML was favoured, as the literature states this abnormality is seen in approximately 1% of cases of AML.
However, the additional positive markers (CD79a and CD22) by flow cytometry confirmed that the diagnosis was B-ALL.
Fig. 2(A) Metaphase and interphase showing KMT2A amplification with >20 signals per cell. (B) Three KMT2A signals (yellow fusion) and KMT2A amplification possibly localised to two intrachromosomal regions. (C) Proposed two KMT2A amplification clones (smaller cells with one localised region of KMT2A amplification beside larger cells with two localised regions of KMT2A amplification) as well as normal cells, raising the possibility of clonal evolution. (D) Three copies of ABL1 (red) and four BCR signals (green). (E) Loss of one TP53 signal (red) and two copies of the centromere 17 probe D17Z1 (green). (F) Doubling clone of the TP53 deletion with two TP53 signals (red) and four copies of D17Z1 (green).
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
and is considered segmental duplication rather than true amplification. Of the five cases reported in adults, three were referred to as therapy related B-ALL
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
They lead to overexpression of the gene and are generally associated with a poor prognosis. In the literature KMT2A amplification has been reported in association with a TP53 gene deletion in AML
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
Duplication or amplification of chromosome band 11q23, including the unrearranged MLL gene, is a recurrent abnormality in therapy-related MDS and AML, and is closely related to mutation of the TP53 gene and to previous therapy with alkylating agents.
FISH for TP53 deletion using the Vysis TP53/CEP17 FISH probe (Abbott Molecular, USA) confirmed that our patient also had a TP53 gene deletion (Fig. 2E). It also suggested that a doubling clone exhibiting TP53 deletion was present (Fig. 2F) which would be in keeping with a complex karyotype. Due to the patient discharging himself from hospital, no sample was available for conventional cytogenetics. Consequently, we could not determine if the amplification was due to extrachromosomal double minutes or intrachromosomal homogeneously staining regions by examining a G-banded karyotype. Additional image capture of over 50,000 images from the directly harvested peripheral blood cells was then performed and found three normal and three abnormal spontaneously dividing metaphases. These abnormal metaphases (Fig. 2A) favour that the amplification was localised to intrachromosomal regions and not due to extrachromosomal double minutes; but was not conclusive as double minutes can tether to chromosomes.
Review of these additional images also further supported the possibility of clonal evolution (Fig. 2C). The first proposed clone was comprised of small cells with 2 KMT2A signals and a smaller number of KMT2A amplification signals localised to a single region, and the proposed second clone consisted of larger cells with 3–4 KMT2A signals and two localised regions of KMT2A amplification (Fig. 2B). If the amplification signals have an intrachromsomal association with 11q, this would assume there are five+ chromosomal copies of this region present in the larger cells. The finding of clonal evolution would be in keeping with the presence of a doubling clone (Fig. 2F) detected using the Vysis TP53/CEP17 FISH probe (Abbott Molecular, USA). Multiplex PCR was negative for TCF3/PBX1, ETV6/RUNX1, KMT2A/AFF1 and BCR/ABL1 gene fusions. Unfortunately no further array or sequencing was performed due to the patient withdrawing from treatment and therefore the presence of other driver mutations
for B-ALL were not assessed. It is generally reported that acute leukaemias with KMT2A rearrangements have a lower additional mutation rate, however we accept the literature is limited in regard to KMT2A amplification in B-ALL.
This is a fascinating case of KMT2A amplification with associated TP53 gene deletion and clonal evolution in adult de novo precursor B-ALL.
Conflicts of interest and sources of funding
The authors state that there are no conflicts of interest to disclose. Funding was provided by the ICPMR-NSWHP ROPP trust fund for research salary and consumables.
References
Swerdlow S.H. Campo E. Harris N.L. World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues. 4th ed. IARC,
Lyon2017
Repeated lineage switches in an elderly case of refractory B-cell acute lymphoblastic leukemia with MLL gene amplification: a case report and literature review.
Duplication or amplification of chromosome band 11q23, including the unrearranged MLL gene, is a recurrent abnormality in therapy-related MDS and AML, and is closely related to mutation of the TP53 gene and to previous therapy with alkylating agents.
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