PF166 RASD1 INHIBITS THE PROLIFERATION OF LEUKEMIA CELLS AND CORRELATES WITH LEUKEMIA RELAPSE IN ADULTS WITH PHILADELPHIA CHROMOSOME‐NEGATIVE B‐CELL ACUTE LYMPHOBLASTIC LEUKEMIA

Abstract

Background:The prognosis of adult B‐cell acute lymphoblastic leukemia (B‐ALL) remains poor due to drug resistance and relapse although the remission rate reaches 80%∼90%. Identifying molecular abnormalities associated with important cellular activities may help to improve the B‐ALL risk stratification and provide potential therapeutic targets. We have previously found that the transcript level of Ras‐related dexamethasone‐induced 1(RASD1) is abnormally increased in adult B‐ALL bone marrow mononuclear cells (BMMNCs) by bioinformatics and quantitative PCR, suggesting that it may be involved in the pathogenesis of B‐ALL. RASD1 is a member of the Ras superfamily. Its abnormal expression has been reported in multiple solid tumors. However, its clinical relevance and biological role in hematopoietic malignancies, especially in B‐ALL remain to be elucidated.Aims:To determine the clinical significance of RASD1 transcript levels in adult B‐ALL and to study its biological role and the potential mechanism in B‐ALL.Methods:Quantitative real‐time polymerase chain reaction(RT‐qPCR) was used to detect RASD1 transcript levels in BMMNCs from 90 newly diagnosed Philadelphia chromosome(Ph)‐negative B‐ALL and 40 healthy donors. The correlation between RASD1 transcript levels and relapse was analyzed. The biological function and potential mechanism were studied with RASD1‐overexpressing and knockdown human B‐ALL cell lines constructed by lentiviral infection. A xeno‐transplant mouse model was used to study the effect of RASD1 overexpression on proliferation in vivo.Results:The transcript levels of RASD1 in bone marrow mononuclear cells from 90 Philadelphia chromosome(Ph)‐negative adult B‐ALL (81.76%, 0.22%∼1824.52%) were significantly higher than those of 40 healthy controls (7.59%, 0.46%∼38.66%, P < 0.001). Patients with low RASD1 transcript levels had a significantly higher 5‐year cumulative incidence of relapse (CIR, 52.0% [37.4%, 66.6%] vs. 36.2% [22.2%,50.2%]; P = 0.013) and a lower relapse‐free survival (RFS, 47.5% [32.9%,62.1%] vs. 63.1% [49.0%,77.2%]; P = 0.012) than patients with high RASD1 transcription levels. Multivariate analysis showed that low RASD1 transcript level was an independent risk factors for CIR and RFS in patients with Ph‐negative B‐ALL (CIR: HR = 3.367[1.668,6.796], P = 0.001; RFS:HR = 2.938 [1.427,6.047], P = 0.003). Functional analysis showed that overexpression of RASD1 in BALL‐1 and Nalm6 cells inhibited cell proliferation, cell cycle progression and survival after exposure to chemotherapeutics including daunorubicin, methotrexate and cytarabine. In contrast, RASD1 knockdown in Sup‐B15 cells showed the opposite effects. The role of RASD1 in inhibiting cell growth was verified in a tumor‐bearing mouse model. Western Blot showed that overexpression of RASD1 caused decreased levels of mTOR and p70S6K phosphorylation, increased expression of LC3BII/I and Beclin‐1, suggesting that mTOR/p70S6K signaling pathway and autophagy may be involved in RASD1‐mediated regulation of cell proliferation.Summary/Conclusion:The transcript levels of RASD1 in BMMNCs from newly diagnosed Ph‐negative adult B‐ALL were significantly higher than those of healthy controls. A low RASD1 transcript level was independently correlated with a higher CIR and a lower RFS. Overexpression of RASD1 inhibited cell proliferation both in vitro and in vivo. mTOR/p70S6K signaling pathway and autophagy may be involved in RASD1‐mediated regulation of proliferation in B‐ALL cells.image