Prognostic stratification improvement by integrating ID1/ID3/IGJ gene expression signature and immunophenotypic profile in adult patients with B-ALL

Abstract

Background: Survival of adults with B-Acute Lymphoblastic Leukemia requires accurate risk stratification of patients in order to provide the appropriate therapy. Contemporary techniques, using clinical and cytogenetic variables are incomplete for prognosis prediction.

Methods: To improve the classification of adult patients diagnosed with B-ALL into prognosis groups, two strategies were examined and combined: the expression of the ID1/ID3/IGJ gene signature by RT-PCR and the immunophenotypic profile of 19 markers proposed in the EuroFlow protocol by Flow Cytometry in bone marrow samples.

Results: Both techniques were correlated to stratify patients into prognostic groups. An inverse relationship between survival and expression of the three-genes signature was observed and an immunophenotypic profile associated with clinical outcome was identified. Markers CD10 and CD20 were correlated with simultaneous overexpression of ID1, ID3 and IGJ. Patients with simultaneous expression of the poor prognosis gene signature and overexpression of CD10 or CD20, had worse Event Free Survival and Overall Survival than patients who had either the poor prognosis gene expression signature or only CD20 or CD10 overexpressed.

Conclusion: By utilizing the combined evaluation of these two immunophenotypic markers along with the poor prognosis gene expression signature, the risk stratification can be significantly strengthened. Further studies including a large number of patients are needed to confirm these findings.

Keywords: B-ALL; Gene-expression; Immunophenotype; Risk-stratification; Survival.

Fig. 1

Gene expression signature predictor of prognosis. a Unsupervised cluster analysis applied to 43 patients with B-ALL according to the expression of our 3-genes signature predictor of prognostic. Cluster analysis discriminates 2 different groups (Green bar good prognosis group and red bar poor prognosis group). Kaplan Meier curves for event-free survival (p = 0.001) b and overall survival (p = 0.001) c according to the presence of 3-genes signature of poor prognosis (red curve) vs patients without the expression profile of poor prognosis (green curve). The expression profile of poor prognosis is defined as the simultaneous overexpression of genes ID1, ID3 and IGJ

Fig. 2

Determination of groups with differential immunophenotypic markers expression. a Unsupervised hierarchical clustering analysis in diagnosis bone marrow of B-LLA patients according to the expression of the evaluated immunophenotypic markers revealed three main groups of patients. Green bar (group 1) corresponds to the group with high rates of complete remission, low number of patients with positive MRD and absence of patients with t (9; 22). While red bar (group 3) represents the group of patients with the highest percentage of induction failures, positive MRD and all patients with t(9; 22). Blue bar (group 2) represents a small group of patients with intermediate outcome compared with the two groups in the extremes. CR: complete remission. MRD: minimal residual disease. b Expression levels of immunophenotypic markers in the two groups formed at the ends of the heatmap with different clinical features (groups 1 and 3). CD45, CD19, CD38 and FSC have lower expression in group 3 than in group 1. CD10 and Kappa have different high expression in group 3 compared to group 1. The differences are significant (*p = <0.05, ***p = <0.0001). Kaplan Meier curves for event-free survival (c) (p = 0.008) and overall survival (d) (p = 0.559) in the two prognostic groups identified by immunophenotypic profile. Note that EFS curves showed statistically significant differences, whereas the curves for OS did not

Fig. 3

Expression levels of genes a ID1 (p = 0.0432), b ID3 (p = 0.024) and c IGJ (p = 0.0399) included in the poor prognosis genetic signature in groups determined by immunophenotypic expression pattern

Fig. 4

a Heatmap and clustering analysis according to immunophenotypic markers expression correlated with poor prognosis genetic signature. Note that at the bottom of heatmap it is shown that patients in green group are those with lower altered expression of ID1, ID3, IGJ genes. While most patients with overexpression of two or more of these genes are included in the red group. Kaplan-Meier curves for event-free survival (p = 0.001) b and overall survival (p = 0.045) c according to the groups defined by the heatmap in a

Fig. 5

Event free survival (a, c) and overall survival (b, d) of 42 patients according to the categorization defined by the presence of the genetic signature and CD10 or CD20 expression. p values correspond to differences between gray and yellow curves