NT5E gene and CD38 protein as potential prognostic biomarkers for childhood B-acute lymphoblastic leukemia

Abstract

The risk stratification of B-acute lymphoblastic leukemia (B-ALL) is based on clinical and biological factors. However, B-ALL has significant biological and clinical heterogeneity and 50% of B-ALL patients do not have defined prognostic markers. In this sense, the identification of new prognostic biomarkers is necessary. Considering different cohorts of childhood B-ALL patients, gene (DPP4/CD38/ENTPD1/NT5E) and protein (CD38/CD39/CD73) expressions of ectonucleotidases were analyzed in silico and ex vivo and the association with prognosis was established. In univariate analyses, expression of NT5E was significantly associated with worse progression-free survival (PFS) in bone marrow (BM) samples. In multivariate analyses, Kaplan-Meier analysis, and log-rank test, higher NT5E expression predicted unfavorable PFS in BM samples. Considering minimal residual disease (MRD), higher levels of cellularity were associated with the high NT5E expression at day 8 of induction therapy. In addition, we observed that white blood cells (WBC) of childhood B-ALL patients had more CD38 compared to the same cell population of healthy donors (HD). In fact, MRD > 0.1% patients had higher CD38 protein expression on WBC in comparison to HD. Noteworthy, we observed higher CD38 expression on WBC than blasts in MRD > 0.1% patients. We suggest that NT5E gene and CD38 protein expression, of the ectonucleotidases family, could provide interesting prognostic biomarkers for childhood B-ALL.

Keywords: CD38 protein; Childhood B-acute lymphoblastic leukemia; Ectonucleotidases; NT5E gene; Prognostic.

Fig. 1

mRNA expression level of genes in BM (N = 198) and PB (N = 91) samples. mRNA expression levels were divided by the distribution’s median. For each gene, patients were labeled as high-expression (red) for expression values above median (values > 50%), while patients were labeled as low-expression (blue) for expression values below median (values ≤ 50%). *p < 0.05, ***p < 0.001. BM, bone marrow; PB, peripheral blood

Fig. 2

Kaplan–Meier curves of childhood B-ALL patients stratified by NT5E (a–b) expression levels. Progression-free survival curve of patients in discovery set with high versus low NT5E expression levels in bone marrow (a) and peripheral blood (b) samples. p-values for significance of difference between high and low expression were calculated using the log-rank test

Fig. 3

Kaplan–Meier curves of childhood B-ALL patients stratified by NT5E (a–b) expression levels. Overall survival curve of patients in discovery set with high versus low NT5E expression levels in bone marrow (a) and peripheral blood (b) samples. p-values for significance of difference between high and low expression were calculated using the log-rank test

Fig. 4

Prevalence of MRD (% of cellularity) at days 8 and 29 of induction therapy in two groups of patients with low and high expression of NT5E in BM samples. From an initial cohort of 198 patients with BM samples, only 130 patients, who had MRD data on both days, were selected. MRD, minimal residual disease

Fig. 5

Gating strategies and cell population definitions (a) and representative plot of CD38 (b and e), CD39 (c and f), and CD73 (d and g) expression on ${\mathrm{SSC}}^{\mathrm{low}}$/${\mathrm{CD}45}^{\mathrm{bright}}$+${\mathrm{SSC}}^{\mathrm{high}}$/${\mathrm{CD}45}^{\dim /\mathrm{bright}}$ cells from HD vs. B-ALL patients (b–d)/B-ALL patients categorized as MRD < 0.1% or MRD > 0.1% (e–g). p values were calculated with non-parametric Mann–Whitney U-test (b–d) and one-way ANOVA test (e–g). *p < 0.05. B-ALL, childhood B acute lymphoblastic leukemia; HD, healthy donors; MRD, minimal residual disease; MFI, mean fluorescence intensity

Fig. 6

Representative summary plot of CD38 (a and d), CD39 (b and e), and CD73 (c and f) expression on ${\mathit{SSC}}^{\mathit{all}}/{CD45}^{dim/bright},{\mathit{SSC}}^{\mathit{low}}$/${CD45}^{\mathit{bright}}$+${\mathit{SSC}}^{\mathit{high}}$/${CD45}^{dim/bright}$, and ${\mathit{SSC}}^{low/int}/{CD45}^{\mathit{dim}}$ cells from B-ALL patients (a–c)/B-ALL patients categorized as MRD < 0.1% or MRD > 0.1% (d–f). p values were obtained with one-way ANOVA (a–c) and two-way ANOVA (d–f) tests. *p < 0.05. MRD, minimal residual disease; MFI, mean fluorescence intensity