Μελέτη του ρόλου της απόπτωσης των νεοπλασματικών κυττάρων στις αιματολογικές κακοήθειες της παιδικής ηλικίας

Abstract

Introduction: Acute lymphoblastic leukemia (ALL) accounts for nearly 1/3 of all pediatric malignancies and 75% of all childhood leukemias. The annual incidence of ALL has been estimated to 30 cases per million, with a peak incidence in children aged two to five years. Progress in the diagnosis with novel molecular techniques, risk classification, and treatment strategy in ALL has led to cure rates that now exceed 80%. However, a significant proportion (20%) of patients fails to respond to therapy, and treatment failure can occur even in patients with favorable prognostic features. It has been suggested that leukemia is characterized by impaired balance between proliferation of blood cells and their capacity to undergo apoptosis. The aim of this study was to assess the expression of the apoptosis-related genes bcl-2 and bax in childhood ALL, both at the time of diagnosis and at remission achieved post induction treatment. In addition, we measured the levels of the apoptotic receptors Fas, FasLigand, and their co-expression on patients’ leukemic cells. To explore the prognostic significance of apoptosis-related genes in childhood ALL, we examined associations between expression levels and established clinical and cytogenetic disease parameters.Materials-Methods: The study included 26 children (eighteen boys, eight girls) with newly diagnosed ALL (twenty-three B-ALL, three T-ALL). The mean age was 7.1 ± 1.2 years, the mean white blood cell count was 27.5 ± 10.6 K/μL and the mean hemoglobin was 9.1 ± 0.6 g/dL. All patients were diagnosed, treated and followed at the Department of Pediatric Hematology-Oncology, University Hospital of Heraklion - Crete, and they received chemotherapy according to the ALL BFM 2000 protocol. There were 34 age-matched children who served as controls (20 children with benign blood diseases -12 with Idiopathic Thrombocytopenic Purpura, 8 with Autoimmune Neutropenias- and 14 children with solid tumors without bone marrow infiltration). Bone marrow specimens were obtained from all children, under informed consent signed by the parents/guardians. Cytogenetic abnormalities were examined with conventional karyotype and FISH. Disease remission following induction therapy was assessed by bone marrow microscopic evaluation and flow cytometry. Measurement of bcl-2 and bax mRNA was performed by quantitative real-time PCR, and membrane expression of Fas and Fas-L was assessed by flow cytometry in bone marrow mononuclear cells, both at diagnosis and at remission following induction chemotherapy.Results: At diagnosis, increased level of the apoptotic bax/bcl-2 ratio was observed in children older than 10 years and with higher white blood cell count. DNA index <1,16 was associated with increased bax/bcl-2 both at diagnosis and at remission, and the del(9p) abnormality with increased bax/bcl-2 at remission. Expression of the apoptotic receptor Fas was significantly higher at remission compared to diagnosis, which might reflect enhanced sensitivity of the leukemic clone to apoptosis and response to treatment. Conclusions: In conclusion, our study highlights the association between the apoptotic bax/bcl-2 ratio with high-risk features in children with ALL, such as older age, white blood cell count, the del(9p) abnormality and DNA index <1.16. The increase in Fas expression once remission has been achieved after induction treatment, could represent a prognostic factor of favorable response to chemotherapy and deserves further investigation. Delineation of the role of apoptosis in pathogenesis and prognosis of pediatric ALL should enable the design of novel targeted therapies.