Diverse asparagine synthetase expression in lymphoid blasts is not relevant to the sensitivity to L-Asparaginase
Abstract
Acute lymphoblastic leukaemia (ALL), the most common haematological malignancy in childhood, is treated by combined chemotherapy, which includes the enzyme L-Asparaginase (L-Asp). The cytotoxic effect of L-Asp consists in its ability to deplete extracellular asparagine and glutamine.
The sensitivity of primary ALL cells to this depletion is traditionally explained by decreased activity of glutamine-dependent enzyme asparagine synthetase (ASNS). Despite the fact that increased ASNS level was indeed shown to be connected with L-Asp resistance, the exact relationship between ASNS expression and L-Asp sensitivity has not been elucidated so far.
The gene expression and ASNS protein content was evaluated in 4 leukemic cell lines: Nalm6 (TEL/PDGFRB[+]); RS4;11 (MLL/AF4[+]); REH (TEL/AML1[+]) and UOCB6 (TEL/AML1[+]). ASNS protein levels reflected ASNS mRNA levels and these correlated negatively with L-Asp sensitivity.
UOCB6 as the most resistant cell line had the highest expression of ASNS, followed by Nalm6, REH and RS4;11. Detection of protein content in primary ALL blasts was not possible due to significantly lower ASNS gene expression compared to cell lines.
Gradient knock-down was performed in 2 ALL cell lines: REH with intermediate basal expression and RS4;11 with very low basal expression. A gradual silencing of ASNS gene in REH cell line led to gradual increase of L-Asp sensitivity.
The reduction of ASNS did not potentiate L-Asp cytotoxicity in RS4;11 cell line. Our data demonstrate that in cells with very low ASNS expression, as shown in primary ALL blasts of various subtypes, the difference in ASNS levels is not relevant to the sensitivity to L-Asp.
We suppose that glutamate dehydrogenase (GDH) is a new player in the response to cytotoxic effect of L-Asp. Silencing of GDH gene in TEL/AML1[+] REH cell line increased sensitivity to L-Asp.
Furthermore, we suggest a relationship between ASNS and GDH based on our observations of increased GDH expression in cells with silenced ASNS gene.