TRANSMEMBRANE ADAPTOR PROTEIN NTAL ENHANCES PROXIMAL SIGNALING AND POTENTIATES CORTICOSTEROID INDUCED APOPTOSIS IN T-ALL

Sborník

Background. The biology of T-lineage acute lymphoblastic leukaemia (T-ALL) is characterized by the pre T-cell receptor (TCR) signaling. Transmembrane adaptor proteins could play an important role in the development of T- ALL. LAT is the adaptor protein essential for signal transmission from TCR. NTAL is an analogue of LAT except for a binding site for PCKgamma. Aims. We analysed the impact of NTAL and LAT expression on biology and treatment response in T-ALL. Methods. NTAL and LAT expression was analysed in diagnostic bone marrow samples of 39 pediatric patients with T-ALL using qRT-PCR and flow cytometry. Jurkat cell line (T-ALL cell line expressing no NTAL) (Jurkat/wt) and derived Jurkat cell line with stable NTAL expression (Jurkat/NTAL+) were used for in-vitro experiments. Cell signalling and cell death after TCR stimulation and after methylprednisolone treatment were analysed using flow cytometry, Western blot and qRTPCR. Results. Leukemic blasts of T-ALL patients responding favourably to initial prednisone treatment had higher levels of NTAL than patients responding unfavourably (p=0.028) whereas LAT expression was stable (p>0.9). This observation was confirmed in in-vitro experiment - after 48 hours of methylprednisolone treatment the percentage of surviving Jurkat/NTAL+ vs. Jurkat/wt cells was 11% vs. 31% (p<0.05). Moreover, Jurkat/NTAL+ cells were more sensitive to TCR induced cell death than Jurkat/wt cells (40.7% vs. 61.0% surviving cells at 24 hours after TCR stimulation, p<0.05). Jurkat/NTAL+ cells showed significantly higher levels of ERK phosphorylation after TCR stimulation (median 1.5 fold, p<0.05) and of CD69 activation marker (90.5% vs. 77.4%, p<0.05) compared to Jurkat/wt, whereas phosphorylation of JNK showed an inverse pattern and phosphorylation of p38 MAPK was stable. The ERK inhibitor U0126 almost completely abrogated TCR induced cell death and reversed sensitizing effect of the NTAL protein to methylprednisolone induced cell death - percentage of surviving Jurkat/NTAL+ and Jurkat/wt cells after 48 hours of coincubation with methylprednisolone and U0126 is 29.3% and 29.9% respectively (p=0.49). Conslusions. We conclude that in our experiment NTAL is a tumor suppressor enhancing ERK phosphorylation of leukemic blasts. ERK molecule is the key protein responsible for increasing cell sensitivity to methylprednisolone induced cell death.This study was supported by grants NS/10473-3, MSM 0021620813 and MSMT NPV 2B06064.

Haematologica, 2012; 97(s1):  256