Quantitative proteomic analysis reveals maturation as a mechanism underlying glucocorticoid resistance in B lineage ALL and JNK inhibitors as a re-sensitising therapy

Glucocorticoids (GC) are pivotal in the treatment of childhood acute lymphoblastic leukaemia (ALL) but resistance is a continuing clinical problem with the underlying mechanisms still unclear. An isobaric tag proteomic approach was used to compare protein profiles of the B lineage ALL GC-sensitive cell line, PreB 697, and its GC-resistant sub-line, R3F9, before and after dexamethasone exposure. Two transcription factors involved in B- cell differentiation, PAX5 and IRF4, were differentially regulated in the PreB 697 compared to the R3F9 cell line in response to GC. PAX5 basal protein expression was less in R3F9 compared to its GC-sensitive parent and was confirmed to be lower in other GC-resistant sub-lines of Pre B697 and was associated with a decreased expression of the PAX5 transcriptional target, CD19. Gene set enrichment analysis of microarray data from the cell lines showed that increasing GC-resistance was associated with differentiation from preB-II to an immature B-lymphocytes stage. GC resistant sub lines were shown to have a higher levels of p-JNK compared to the parent line and JNK inhibition caused re-sensitisation to GC. Reduced CD19 levels accompanying GC resistance was also apparent in some clinical samples, with high levels of MRD persisting after GC containing induction chemotherapy. Thus, quantitative proteomic analysis reveals a role for PAX5 and maturation as a recurrent mechanism underlying glucocorticoid resistance in ALL and identifies JNK inhibitors as a possible re-sensitising therapy. Gene expresion profiling of GC-sensitive and resistant precursor B-ALL cells. Gene set enrichment analysis (GSEA) was used to analyse the state of differentiation of GC-resistant sub-lines and genes were ranked according to the correlation of gene expression with dexamethasone IC50 values for the individual sub-lines.