B-cell lymphoblastic leukemia/lymphoma with mutated IKZF1 N159Y: clinical and genetic features of an emerging entity

IKZF1 (IKAROS family zinc finger 1) alterations, most commonly partial or full gene deletions, occur in about 15% of pediatric and 40% of adult B-lymphoblastic leukemia/ lymphoma (B-ALL) cases and are considered a poor prognostic marker.¹ However, IKZF1 somatic point mutations are uncommon. B-ALL with mutated IKZF1 N159Y (p.Asn159Tyr), classified as a new entity per the International Consensus Classification of Myeloid Neoplasms and Acute Leukemia, represents a rare subtype of B-ALL with an estimated incidence of <1% and only a few reported cases.^2-5 This is the first comprehensive study that provides the clinical, immunophenotypic, and genetic features. We found that this rare entity occurs in more than half of pediatric patients, typically presenting with an abnormal karyotype, a gain of chromosome 21, absence of a fusion transcript, and a few additional genetic abnormalities.

We searched our pathology archives, collaborated with nearby colleagues and performed a thorough literature review, identifying in total of 13 patients with an IKZF1 N159Y mutation. Overall, two patients were from our institution, one patient was from the University of Chicago, and ten patients were documented in the literature, with some overlap between two studies and a single case from a different study.^3-5 Clinical and demographic characteristics of these 13 patients were reviewed. The genetic characteristics (conventional cytogenetic, fluorescence in situ hybridization [FISH] and comprehensive next-generation sequencing [NGS]) which included analysis of RNA, DNA and copy number variants (CNV) were analyzed. Classification of variants identified through NGS was performed per the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) criteria. Positive minimal residual disease in all patient samples collected at the end of induction was defined as 0.01% or higher by flow cytometry analysis except for patient #10 where polymerase chain reaction for IgG rearrangement was performed. This study was conducted in compliance with the ethical principles of the Declaration of Helsinki and our institutional review board policy.

Although B-ALL with mutated IKZF1 N159Y occurred in patients from 2 to 45 years of age with no clear predilection for males or females, more than half of the cases occurred in pediatric patients (61.5%, 8/13) (Figure 1A; Table 1). The pediatric patient group contained standard-risk (SR) and high-risk (HR) patients (4 SR and 3 HR) (Table 1).

Interestingly, patients often presented with white blood cell counts within the normal range (2.8-16.7x10⁹/L) (67%, 8/12) (Table 1). However, none of the cases identified in the literature provided details regarding clinical presentation and blast immunophenotype. The three patients (patients #11-12 from our institution and patient #13 from University of Chicago) described in the current study presented with anemia (range, 6.3-7.3 g/dL) and had no evidence of clinical nervous system (CNS) involvement by leukemic cells at the time of diagnosis. Additionally, all three patients exhibited similar blast morphology and immunophenotype. The blasts expressed CD10 (bright), CD19, CD22, CD34, CD38 (dimmer than normal to negative), CD79a, TdT; and were negative for CD20, CD33 and other myeloid and T-cell markers. This is the first report highlighting the immunophenotype by flow cytometric analysis for all three B-ALL patients with IKZF1 N159Y. Notably, all patients who had reached the end of induction at the time of review were negative for minimal residual disease (100%, 8/8), and had reached complete remission (100%, 4/4) (Figure 1A; Table 1).

Figure 1.Demographic, clinical, and genetic characteristics of B-lymphoblastic leukemia/lymphoma with mutated IKZF1 N159Y (N=13). (A) Demographic and clinical characteristics. (B) Genetic abnormalities. AYA: adolescent and young adults; WBC: white blood cell count; MRD: minimal residual disease; Chr.: chromosome.

Table 1.Demographic and clinical data.

Chromosome analysis revealed that most patients harbored an abnormal karyotype (90%, 9/10) while 83% of patients (10/12) had a gain of chromosome 21 as determined by cytogenetic analysis and/or copy number variations from exome sequencing (Table 2). Notably, additional mutations were uncommon, with pathogenic variants classified as Tier II, variants of potential clinical significance in genes involved in RAS signaling pathway (KRAS and FLT3; 15%, 2/13) (Table 2). Variants classified as Tier III, variants of uncertain clinical significance (VUS) observed in this study were either not reported in other databases (ETV6, KMT2D), previously documented to be germline variants per the ClinVar database (RUNX1) or classified as both germline and somatic variants per ClinVar and Cosmic databases (IL7R and NOTCH1) (Table 2). Computational predictive tools suggest that variants of uncertain significance (KMT2D, RUNX1, IL7R and NOTCH1) may not impact protein. Although we cannot know with certainty if these variants are somatic or germline due to the analysis of only neoplastic samples, the allele frequency and available data are more in favor for RUNX1, IL7R and NOTCH1 VUS variants being of germline origin. Fusion transcripts were not detected by NGS or reported in the literature (0%, 0/13) (Table 2). Two patients, #6 and #11 showed chromosomal translocations that were detected by chromosome analysis t(6;21)(p23;q11.2) and t(14;21) (q21;q22), respectively. However, it is currently unknown if these translocations generate functional fusion products as they have neither been reported in the literature nor observed with RNA analysis.

This study identified that B-ALL with IKZF1 N159Y is strongly associated with a gain of chromosome 21. Trisomy of chromosome 21 in the constitutional setting is diagnostic of Down syndrome and is associated with an increased risk of acute lymphoblastic leukemia.⁶ Approximately 35% of Down syndrome-associated B-ALL show an IKZF1 deletion.⁷ However, the gain of chromosome 21 in patients with B-ALL with IKZF1 N159Y is a somatic mutation and thus is not associated with Down syndrome. Furthermore, unlike other IKZF1 deletions, commonly associated with BCR-ABL1-like B-ALL, patients with B-ALL with mutated IKZF1 N159Y appear to have very few additional genetic abnormalities. Specifically, no fusion transcripts were detected by NGS or reported in the literature.^3-5

Although this study is limited in the number of patients due to the rarity of this emerging entity being less than 1% of B-ALL, patients with B-ALL with mutated IKZF1 N159Y, in this small cohort, had a good response to treatment with 100% (8/8) being negative for minimal residual disease at the end of induction and remaining in remission after diagnosis (Table 1). In contrast, many studies have identified IKZF1 deletions as an adverse prognostic predictor, commonly associated with BCR-ABL1-like B-ALL.

Table 2.Genetic abnormalities (cytogenetic and molecular analysis).

IKZF1 is located on chromosome 7p12.2 and consists of eight exons. Most IKZF1 deletions occur in exons 4-7, resulting in expression of the IK6 isoform, a dominant-negative form of IKAROS, that lacks the DNA-binding domain. IKZF1 deletions lead to a dominant-negative IKAROS effect, in part by mislocalizing IKZF1 from the nucleus to the cytoplasm.^8,9

Exons 4 to 6 are thought to be essential for maintaining IKZF1 tumor suppressor function. The IKZF1 N159Y missense variant is located in exon 5, in the DNA-binding domain of IKZF1. IKZF1 missense variants are commonly located at or near residues known to be critical for DNA binding.¹⁰ Previous studies have shown that the IKZF1 N159Y variant disrupts IKZF1 function, resulting in distinctive nuclear mislocalization and the induction of aberrant intercellular adhesion characteristic of many IKZF1 alterations.⁹ While IKZF1 missense variants have a damaging effect on the protein, the missense variants may be less deleterious when compared to partial or whole gene deletions. Also, the deleterious effect of IKZF1 alterations may be confounded by other co-occurring genomic aberrations. For example, patients with co-occurring IKZF1 and ERG deletion, commonly observed in B-ALL with DUX4 rearrangement tend to have better clinical outcomes.^11-13 In contrast, IKZF1 deletions co-occurring with deletions in CDKN2A, CDKN2B, PAX5, or the pseudoautosomal region PAR1 in the absence of ERG deletion are thought to be adverse prognostic indicators.^14-15 Importantly, B-ALL with mutated IKZF1 N159Y missense mutation displays a strikingly different gene expression profile compared to other B-ALL cases, including other IKZF1-altered cases.^3-4 B-ALL with mutated IKZF1 N159Y exhibited upregulation of different genes from those observed to be upregulated in patients with other IKZF1 alterations and also showed downregulation of genes involved in B-cell receptor signaling and JAK-STAT signaling, such as FLT3 and STAT5A.^3-4

In conclusion, IKZF1 N159Y mutation is a rare recurrent abnormality in B-ALL. This unique entity occurs more commonly in pediatric patients and has a strong association with an abnormal karyotype and acquired gain of chromosome 21. Additionally, patients with B-ALL with IKZF1 N159Y mutation have no detectable fusion transcripts, have very few additional genetic abnormalities, and based on available data, all patients who had reached the end of induction were negative for minimal residual disease.

Footnotes

• Received September 17, 2024
• Accepted February 17, 2025

Correspondence

A. I. RICHARDSON - airichardson@luriechildrens.org/aida.richardson@northwestern.edu

Disclosures

No conflict of interests to disclose.

Contributions

Acknowledgments

Revised manuscript language was edited by Anne Sally Davis through Eloquenti.