iAMPlified gene expression offers new insights in B cell precursor leukemia subtype

Acute lymphoblastic leukemia (ALL) is the most prevalent childhood cancer and represents approximately 25% of cancer diagnoses among children younger than 15 years. About 6,000 children and adolescents are diagnosed with ALL each year in the US and there is a gradual increase in the incidence¹. Although current treatments, including systemic chemotherapy and radiation, show relatively high response rates (up to 80%), they can be burdensome with high risk of secondary malignancies. Relapsed and refractory patients have few treatment options. Immunotherapy using engineered T cells appears promising for some patients, but these experimental therapies exhibit some toxicity. Thus, ALL represents a highly unmet need due to the toxicity of current therapy, the absence of targeted therapies and the aggressive relapsed cases of this disease. ALL patients present with a suppression of several blood lineages including thrombocytopenia that causes bruising, anemia that can cause fatigue and neutropenia leading to infections. Metastasis to the central nervous system and testicles is also common.

Although about 80% of children ALL patients respond to chemotherapy (standard risk), the patients that do not respond (refractory disease) or relapse post chemotherapy (high risk) have limited possibility for survival. Clinical, immunophenotypic and cytogenetic markers have been used for the disease prognosis. For instance, high blast number at diagnosis, persistence of blasts during or fast reemergence during chemotherapy (manifested as the detection of clones or minimal residual disease, MRD), T-cell immune-phenotype and lower number of chromosomes (hypodiploidy) associate with poor disease prognosis.

ALL patients present with about 20 genetic alteration events at the time of diagnosis¹. Translocations frequently associate with prognosis. For instance, the presence of the Philadelphia chromosome t(9;22)(q34;q11) associates with poor prognosis, whereas RUNX fusions such as t(12;21)(p13;q22)/ETV6-RUNX1 associate with good prognosis. Intrachromosomal amplification of chromosome 21 (iAMP21) exists in a 2% of pediatric cases of B-cell precursor ALL (BCP-ALL), it was first detected in 2003^2,3, is considered a primary event in this disease subtype⁴, and it is associated with a median patient age of 9 years at the time of diagnosis and intermediate to poor prognosis. Indeed, these cases have poor prognosis when treated with standard chemotherapy whereas respond well to aggressive chemotherapy protocols⁵.

Although iAMP21 consists of heterogeneous spectrum of alterations including multiple regions of gain, amplification, inversion, and deletion, detection of four or more copies of RUNX1 on a single abnormal chromosome 21 (a total of 5 or more RUNX1 signals per cell) using FISH is a means of detection of iAMP21, as RUNX1 belongs to the commonly amplified 5.1 Mb region of chromosome 21 from 32.8 to 37.9 M. iAMP21 can rarely co-exist with BCR-ABL1, or ETV6-RUNX1 translocations. Nevertheless, the role of RUNX1 in this disease subtype is not hitherto characterized and surprisingly RUNX1 expression is not particularly elevated in iAMP21 cases compared to other cases with chromosome 21 abnormalities. This disease subtype is substantially understudied.

In the current volume of Leukemia & Lymphoma, Barbany and colleagues attempted to describe expression profile of iAMP21 cases, taking into consideration that the genomic landscape of this subtype of B-cell leukemia has been previously characterized^4,6,7. The group performed RNA sequencing and copy-number alterations analysis in 12 iAMP21 samples to discover that about half of the genes in the amplified region are over-expressed and they identified three genes as the most highly expressed in this disease subtype: the dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) kinase^8–12, the chromatin modulator Chromatin Assembly Factor 1 Subunit A (CHAF1)^13–15 as well as the RNA binding factors SON (or Negative Regulatory Element-Binding Protein¹⁶) and the Zinc Finger CW-Type Coiled-Coil Domain Protein 3 (MORC3)¹⁷. The authors confirmed that DYRK1A, CHAF1 and SON present with significantly higher expression in iAMP21 compared to other BCP-ALL subtypes. Interestingly, expression levels for those genes do not associate with DNA methylation levels in the corresponding loci. The authors also describe translocations between RUNX1-DYRK1A that do not affect the expression of RUNX1 and do not further upregulate DYRK1 at least in a statistically significant way. A fusion transcript joining exon 2 of RUNX1 with one of the first exons of DYRK1A could be confirmed in one of the cases. Finally, although the group described copy-number alterations in relapsed compared to diagnosis disease, there are no available gene expression data from relapse cases in the study that could potentially help interpret mechanisms of relapse.

The findings pave the way for further research in iAMP21 B-ALL’ SON and MPRC3, for instance, might control RNA biology and splicing, that has been shown be deregulated in leukemia¹⁸. Moreover, DYRK1A and CHAF1B have been associated with poor prognosis in acute lymphoblastic leukemia and myeloid disorders. DYRK1A, a gene belonging to the Down syndrome critical region, a set of critical 33 genes in trisomia 21, can promote megakaryocytic leukemia via suppression of NFAT activity¹⁹ and is essential for lymphoid, but not myeloid cell development mainly via phosphorylation of cyclin D3¹² and it can thus control the balance between proliferation and differentiation of lymphoid cells. To this end, inhibition of DYRK1A can be a therapeutic modality in B-cell leukemia.

CHAF1B, in turn, regulates DNA replication and repair as part of the trimeric chromatin assembly complex 1 (CAF1) consisting of p48, CHAF1B (p60), and CHAF1A (p150) that deposits histones into the newly replicated DNA^13,14. CHAF1B expression associates with poor prognosis in cancer²⁰. As CAF1 complex controls DNA repair and could contribute to response to chemotherapy. To this end, gene expression data from relapsed tumors will be critical in further dissecting mechanistic contributions from the genes described in the current study. It was shown that CHAF1B also regulates gene expression in acute myeloid leukemia via binding to chromatin sites with motifs for transcription factors that control terminal differentiation, such as FLI1 and CEBPA¹⁵. CHAF1 is believed to promote leukemia via blocking the binding of these sequence specific transcription factors that guide normal myeloid differentiation.

Barbany and colleagues showed that two of the FLI1 targets, HOXA10 and RB1, are downregulated in iAMP21 BCP leukemia. Nevertheless, further research is required to dissect pathways affected by CHAF1B and DYRK1A activities and whether the two proteins converge down to the same or similar pathways in iAMP21. In a recent study, Harrison and colleagues identified that the RAS pathway is activated in iAMP21 samples associated with sensitivity to MEK1/2 inhibitor, selumetinib²¹. The role of the genes identified in the current study in disease as potential interactants with the RAS pathway needs to be further explored.

In conclusion, the current is an interesting study that disentangles the genetic alterations from the gene expression changes and it might pave the way for further mechanistic studies in iAMP21 on the roles of DYRK1A, CHAF1B and SON towards leukemia promotion and their interactions with other oncogenic pathways in this disease.