Expression of LAT and PAG Adaptor Molecules in Blasts of Childhood Acute Lymphoblastic Leukaemias and in Normal B- and T-Precursors

Despite the progress in leukaemia research, genetic background of some acute lymphoblastic leukaemias (ALL) is unknown. In the aetiopathogenesis of ALL the transmembrane adaptor proteins (including LAT and PAG) could play an important role as these proteins are key mediators of signalization cascades in lymphocytes.
We analysed expression of PAG and LAT at the mRNA level by real-time quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and at the protein level by flow cytometry. In our study we assessed the expression levels in 10 patients with T-ALL and in 10 patients with B-cell precursor ALL (BCP-ALL). All analysed samples had a high percentage of malignant blasts (70-97%, median 91%). Sorted and unsorted bone marrow/peripheral blood samples of healthy volunteer donors were used as controls. To determine the PAG and LAT expression levels in normal immature thymocytes we analysed also sorted cells from the normal thymuses taken from children undergoing heart surgery. We used various differentiation markers of both B- (CD10, CD19, and CD20) and T-lineage (CD1a, CD3, CD4, CD7, CD8) for sorting to compare the leukaemic samples with the most adequate control in terms of maturation status of the cells. Informed consents were obtained from all volunteer donors or their guardians.
Our data show that the expression of PAG at both protein and mRNA level during the physiological B-cell maturation is decreasing. On the contrary, PAG expression during the T-cell development and also the levels of LAT during the B- and T-cell maturation are increasing, however, the levels of LAT mRNA in B-precursors are very weak.
In childhood BCP-ALL the expression of PAG on protein and mRNA level reflects the situation in corresponding non-malignant precursors with the exception of the TEL/AML1 positive leukaemias that show tendency to the overexpression of PAG. In the T lineage ALL we found a significantly lower expression of PAG and higher expression of LAT mRNA ((p<0.0001 and p=0.03, respectively, for mature T-ALL) compared to sorted thymocytes with corresponding immunophenotype. The LAT protein in BCP-ALL blasts is expressed very weakly or there is no expression at all. Also the levels of LAT mRNA are even lower than in the physiological BCP counterparts.
We present the first study assessing changes in the expression of two important adaptor molecules at both mRNA and protein levels during the physiological maturation of lymphocyte precursors. Moreover, we demonstrate changes in the pattern of expression in childhood ALL and suggest that the aberrant transmission or processing of signal through the immunoreceptors of lymphoid precursors might play an important role in the aetiopathogenesis of the disease.
Supported by grants GAUK 50/2005, MSMT 21620813, IGA NR9108-3 and MSMT 2B06064.