Refractory/relapsed ALL (RR-ALL) is the first priority issue to clinicians and translational researchers. Multiple layers of factors, e.g., somatic genomic lesions, inherited variation, micro-environment, and acquired mutations, played important roles in RR-ALL. Recent studies showed that FOXO1 was a predominant transcription factor in B-lineage-restricted progenitor cells. Pre-BCR signaling activation can suppress FOXO1 transcription activity and subsequent B-ALL cells maintenance. However, no report has been focus on the role of FOXO1 in B-ALL prognosis.
In this study, we identified a novel FOXO1 fusion gene in an induction failure B-ALL patient, namely MEIS1-FOXO1. FOXO1 gene belongs to the forkhead family of transcription factors, which play roles in myogenic growth and differentiation, cancer development, and therapy [11–14]. Translocation of FOXO1 with PAX3 and PAX7 has been reported in pediatric alveolar rhabdomyosarcoma [8, 15]. In the meanwhile, two cases of FOXO1 fusions with unknown genes have been identified in BCP-ALL , while the role of FOXO1 in B-ALL remains clarified. Using an IL3-dependent growth mouse hematopoietic progenitor cell line Ba/F3, we found that MEIS1-FOXO1 in combination with NRASG12D efficiently induced Ba/F3 cells IL3-indepdent growth and transformation in a faster fashion as compared to NRASG12D alone, suggesting its oncogenic effect. Also we observed that MEIS1-FOXO1 potentiated S-phase entry.
Robert and Mark et al have demonstrated that Foxo1 is a key regulator B cells development, in which Foxo1 inactivation cause differentiation blockage at the pro-B cell stage, suggesting Foxo1 function as a tumor suppressor in mouse[16, 17]. To initially probe the role of MEIS1 and FOXO1 in hematopoiesis and leukemogenesis, especially in B cells differentiation, we first analyzed their expression during hematopoiesis and B-ALL [18–20]. Interestingly, gradual up-regulation of FOXO1 expression was observed during B cells differentiation while no characteristic pattern was identified in MEIS1, suggesting the important role of FOXO1 in B cells development. In combination with the result that the FOXO1 transcription activity was almost completely abolished in the MEIS1-FOXO1 protein, while it MEIS1 transcription activity was normal. Our novel fusion gene finding and its FOXO1 loss of function supported FOXO1 also functioned as a tumor suppressor.
The FOXO1 transactivation activity was completely abolished in MEIS1-FOXO1 as compared to wild-type FOXO1, suggesting that the FOXO1 was totally loss of function in this induction failure patient. FOXO1 has been reported as a predominant transcription factor in B-lineage-restricted progenitor cells [16, 17]. Pre-BCR signaling activation can suppress FOXO1 transcription activity and subsequent B-ALL cells maintenance [21, 22], however, the role of FOXO1 in B-ALL risk stratification and relapse remains unclear.
To address this question, we used sets of dataset to associate the FOXO1 expression with risk stratification. The FOXO1 was highest expressed in ETV6-RUNX1 fusion cases (a well-known excellent prognosis group) while lowest expressed in infantile leukemia, this pattern was confirmed by different cohort studies . Interestingly, we also identified the correlation between lower FOXO1 expression and higher minimal residual disease (MRD) burden post induction therapy (P = 3.8 × 10− 4). All these evidences suggested that lower FOXO1 expression might be one of the independent prognostic factor in childhood B-ALL.
Through real-time quantitative PCR assay, we found that the FOXO1 transcription was significantly lower in relapsed samples, suggesting that FOXO1 loss of function might be an important factor in B-ALL relapse. We performed this association analysis again using serial datasets and reached the same pattern. To gain a more convincing evidence, we tested the FOXO1 transcription level among the matched diagnosis-relapse paired samples to further confirm our findings. Using our study cohort, PCGP data and GSE28642, we uniformly confirmed that FOXO1 was significantly lower expressed in relapse samples than their primary diagnosis samples. Question still remained that how lower FOXO1 expression did contribute to higher MRD level and relapse, we found that lower FOXO1 expression was significantly correlated with glucocorticoid resistance, a very key component in the ALL therapy.