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The effect of aerobic glycolysis remains elusive in pediatric acute T lymphoblastic leukemia (T-ALL). Increasing evidence has revealed that dysregulation of deubiquitination is involved in glycolysis, by targeting glycolytic rate-limiting enzymes. Here, we demonstrated that upregulated deubiquitinase USP1 expression correlated with poor prognosis in pediatric primary T-ALL samples. USP1 depletion abolished cellular proliferation and attenuated glycolytic metabolism. In vivo experiments showed that USP1 suppression decreased leukemia progression in nude mice. Inhibition of USP1 caused a decrease of both mRNA and protein levels in LDHA, a critical glycolytic enzyme. Moreover, USP1 interacted with and deubiquitinated PLK1, a critical regulator of glycolysis. Overexpression of USP1 with upregulated PLK1 was observed in most T-ALL patient samples. In addition, PLK1 inhibition reduced LDHA expression and abrogated the USP1-mediated increase of cell proliferation and lactate level. Ectopic expression of LDHA can rescue the suppressive effect of USP1 silencing on cell growth and lactate production. Pharmacological inhibition of USP1 by ML323 exhibited cell cytotoxicity in human T-ALL cells. Taken together, our results demonstrated that USP1 may be a promising therapeutic target in pediatric T-ALL.

Purpose: Classical introduction therapy of etoposide combined with cytarabine and daunorubicin (DAE) is commonly applied in childhood acute myeloid leukemia (AML), but etoposide has an increasing risk of secondary cancer. In this study the non-inferiority effect of homoharringtonine (H) versus Etoposide was compared in induction phase for Chinese childhood AML treated by CCLG-AML 2015 protocol. Patients and Methods: The total of 818 childhood AML patients (median age of 80 months; range from 1 to 193 months) from CCLG-AML 2015 study group (40 centers) were randomly allocated to two induction arms of DAE and DAH. During the course of induction I, 467 patients in DAE group received daunorubicin and cytarabine (DA) plus etoposide (D: 40 mg/m2 per day on days 1, 3 and 5; A: 100 mg/m2 every 12 hours from day 1 to 7; E: 100 mg/m2 per day from days 1 to 5), and 351 patients in DAH group received the same DA does plus homoharringtonine ( H: 3 mg/m2 per day from days 1 to 5). During the course of induction II, Idarubicin (10 mg/m2 per day on days 1, 3 and 5) was used to instead of daunorubicin, and patients accepted corresponding IAE or IAH treatment. All patients were divided into standard, intermediate or high risk group (SR, IR or HR group) according to CCLG-AML 2015 regimen (table 1). They were assessed by bone marrow (BM) aspiration and morphologically defined complete remission (CR: blasts ≤5%), partial remission (PR: blasts between 6~19%), or non-remission (NR: blasts ≥20%) on days 28 of induction. Results: DAH/IAH group showed non-inferiority for remission rates both in induction Ⅰ (DAE 70.2% vs DAH 76.6%, P = 0.041) and induction Ⅱ (IAE 79.4% vs IAH 87.7%, P = 0.016). Total CR rate at end of induction Ⅰ reached 73.0% and it didn't differ between DAE and DAH group for IR or HR group (IR group: DAE, 73.9% vs. DAH, 77.3%, P = 0.529; HR group: DAE, 53.9% vs. DAH, 62.6%, P = 0.128). But for SR group, CR rate of DAH group is significantly higher than DAE group (DAE, 85.1% vs. DAH, 95.1%, P = 0.013). It has similar results after induction Ⅱ. Total CR rate reached 83.1% and all patients has almost gained CR/PR for SR or IR group, only 2 patients still couldn't obtain remission. There was no significant difference in SR or IR group between two arms, but for HR group, CR rate significantly increased in those who accepted IAH chemotherapy (SR group: IAE, 91.2% vs. IAH, 95.0%, P = 0.398; IR group: IAE, 87.1% vs. IAH, 92.5%, P = 0.275; HR group: IAE, 66.1% vs. IAH, 78.8%, P = 0.050). Conclusion: Homoharringtonine is an effective cytotoxic drug and DAH regimen showed non-inferiority induction effect compared with classical DAE regimen in childhood AML, especially for patients of standard risk group. Disclosures No relevant conflicts of interest to declare.

Objective Intravenous arsenic trioxide (ATO) and all-trans retinoic acid have become the front-line treatments for patients with acute promyelocytic leukemia (APL). Realgar-Indigo naturalis formula (RIF), an oral arsenic drug, not only shows a clinical efficacy comparable to ATO but also promotes incorporating an outpatient postremission therapy model into clinical practice for both low-risk and high-risk APL patients in China. However, the safety of ATO/RIF used in children with APL is unknown. To assess the safety of arsenic (ATO/RIF) administrated in children with APL, we designed the study. Methods Children with newly diagnosed APL treated with CCLG-APL2016 protocol (ChiCTR-OIN-17011227) in Beijing Children's Hospital from July 2016 to December 2018 were eligible for the study. The arsenic concentrations in different tissues, including plasma, urine, hair and nail, were measured at 10 time points: before ATO/RIF(D0), on the 7th, 14th and 28th day of ATO/RIF administration(D7, D14, D28), on the end of consolidation therapy (Con), during the 10 weeks of maintenance therapy (W10), on the cessation of therapy, after six months, one year and 1.5 years without arsenic administration. Adverse reactions were observed to evaluate the safety of arsenic received in children with APL. Results Nineteen patients with newly diagnosed APL were enrolled, including 9 boys and 7 girls with an onset average age of (10.9±3.7) years. There were 14 patients in low-risk group (white cell count Conclusions Through the detection of arsenic concentration in different periods and different tissues, it was found that the plasma arsenic concentration could be maintained within the effective concentration range in each treatment stage, and the arsenic concentration in plasma and hair gradually decreased to normal after six months off arsenic. Urine and nail arsenic went down to normal after 1 year off arsenic. Up to now, with a longest follow-up period of 34.7 months and the mean follow-up time of 19.6 months, the short-term response of arsenic disappeared after symptomatic therapy or arsenic reduction, and no chronic side effects of arsenic were observed. Therefore, the use of ATO/RIF in children with APL is safe, but it still needs long-term follow-up. Disclosures No relevant conflicts of interest to declare.

Purpose DNMT3A mutation has been found in approximately 20% of adult acute myeloid leukemia (AML) patients and in 0~1.4% of children with AML, and the hotspots of mutations are mainly located in the catalytic methyltransferase domain, thereinto, mutation R882 accounts for 60%. Although the prognosis of adult patients with DNMT3A mutations is dismal, the prognosis of DNMT3A mutation in childhood AML is still unclear. Here, we tried to determine the incidence and prognostic significance of DNMT3A mutations in Chinese childhood AML. Methods We detected the mutations in DNMT3A exon 23 by PCR and direct sequencing in 342 children with AML (0~16 years old) from January in 2005 to June in 2013, treated on BCH-2003 AML protocol. The correlation of DNMT3A mutations with clinical characteristics, fusion genes, other molecular anomalies (FLT3-ITD, NPM1, C-KIT and WT1 mutations), and treatment outcome were analyzed. Results DNMT3A mutations were detected in 4 out of 342 (1.2%) patients. The mutations in coding sequences included S892S, V912A, R885G, and Q886R. Furthermore, there were two intronic mutations (c.2598-15C>T and c.2739+55A>C) found in one patient. No association of DNMT3A mutations with common clinical features was found. Two patients with DNMT3A mutations died of relapse or complications during treatment, one patient gave up treatment due to introduction failure in day33. Only one patient achieved continuous complete remission. Conclusions DNMT3A mutations were rare in Chinese children with AML. The mutation positions were different from the hotspots reported in adult AML. DNMT3A mutations may have adverse impact on prognosis of children with AML. Disclosures No relevant conflicts of interest to declare.

BackgroundMethotrexate (MTX) is a key chemotherapeutic agent in the treatment of pediatric acute lymphoblastic leukemia (ALL). Nevertheless, MTX can cause severe adverse effects and toxicities. The aim of the present study was to identify genetic polymorphisms in candidate genes of the MTX pathway associated with MTX pharmacokinetics, toxicity, and outcome in ALL in China. MethodsThree hundred and twenty-two Chinese children with ALL in the standard-risk and medium-risk treatment branches from the Beijing Children's Hospital-2003 and Chinese Childhood Leukemia Group-2008 protocols were enrolled in this study. Sequenom MassARRAY was used to genotype 12 single nucleotide polymorphisms (SNPs) in 4 candidate genes of the MTX/folate pathway. A total of 1268 high-dose MTX (HD-MTX) courses were analyzed. The plasma MTX levels were evaluated at 48 h after the first dose of HD-MTX infusion. Oral mucositis during the consolidation therapy period was recorded. Results No polymorphism was associated with clinical features, prednisone response, and minimal residual disease (MRD) at day 33 and 78. Long-term outcome was better in SLCO1B1 rs4149056 T and TC allele carriers than patients with C allele (5-year RFS 92.3±1.6% vs. 27.8±23.2%,P As to MTX pharmacokinetics, ABCB1 rs1128503 showed a significant association with serum MTX levels (P=0.004). SNPs (rs3788200, rs1131596, rs1051266) of the SLC19A1 gene were also associated with the plasma levels of MTX (P=0.003, 0.004, and 0.003, respectively). No association was found between oral mucositis with any polymorphism. Conclusions Genetic variations substantially influence the kinetics and response to HD-MTX therapy in childhood ALL. Disclosures No relevant conflicts of interest to declare.

Abstract 4811 Background: Mixed phenotype acute leukemia (MPAL) is a rare type of acute leukemia with a poor clinical outcome, defined by the blasts express antigens of more than one lineage. MPAL was also named as mixed lineage leukemia, bilineal leukemia, biphenotype leukemia historically. The properly diagnosis of MPAL is crucial for the treatment. The scoring system proposed by European Group for Immunological Characterization of Leukemia (EGIL) in 1995 was the first standard criteria to diagnose MPAL. In 2008, WHO proposed a new scoring criteria to define MPAL. But both of these two systems have their limitation. Methods: We retrospectively analyzed 41 pediatric patients with mixed phenotype acute leukemia from 2000 to 2010 according to EGIL scoring system. All the 41 patients were reclassified according the 2008 WHO classification. Diagnosis, cytogenetics, immunophenotype, treatment and prognosis were analyzed. Results: There were 25 male and 16 female with the mean age of 8 years old. According to EGIL scoring system, the phenotype included T/myeloid (n=16), B/myeloid (n=13), T/B(n=11) and T/B/AML(n=1), respectively. Among the patients, there were 4 MPAL with t(9;22)(q34;q11), 5 with t(12;21)(p13;q22), 1 with inv(16)(p13;q22) and 1 with t(9;9). The overall complete remission (CR) rate was 47% treated by ALL regimen (n=19) and 17% by the AML+ALL combined therapy (n=6). MPAL with t(9;22)(q34;q11) showed poor response (4/4). MPAL with t(12;21)(p13;q22) had a good response to the treatment(4/5). Based on the 2008 WHO criteria, 24 of the 41 MPAL patients were reclassified as MPAL, including T/myeloid type(n=11), B/myeloid type(n=6), T/B(n=7). Three MPAL patients with t(12;21)(p13;q22) diagnosed by EGIL system were excluded by 2008 WHO criteria. Conclusion: Our results suggest that MPAL is a poor outcome disease requiring more accurate classification based on genetic and gene classification. MPAL with t(9;22)(q34;q11) is a predictor of poor outcome. MPAL with t(12;21)(p13;q22) could be treated only by ALL regimen. MPAL with both lymphoid and myeloid markers appears to achieve a better CR rate if treated by ALL regimens alone than treated by ALL+AML combined regimens. Further studies are needed to determine a standard diagnosis and treatment approach for MAPL. Disclosures: Zheng: Beijing Health System High-level Technical Personel Plan: Research Funding.

To investigate the anxiety and depression of parents whose children suffered from leukemia and to develop a set of theoretical guidance for later psychologic intervention, parents of leukemia children treated in Beijing Children’s Hospital (BCH) from March of 2004 to March of 2006 were evaluated with Family Environment Scale (FES) and Hospital Anxiety and Depression (HAD) questionnaires. They were tested in three phases relating to three groups: parents of newly-diagnosed-children group, parents with or without psychological intervention groups. Parents in intervention group were given individual psychotherapy, group therapy or systemic family therapy. A total of 353 questionnaires were analyzed, including 138 in newly-diagnosed-children group, 103 in intervention group and 112 in non-intervention group. Results showed: The family environment scores of expressiveness, independence, achievement, cultural, recreational orientations and organization were significantly lower, but the scores of conflict, ethics religion and control were much higher in parents of newly-diagnosed-children group than the normative population. For example, the scores of expressiveness and conflict were 4.9 and 2.3 in normative population, but the scores were 2.3 and 3.9 in parents of leukemia children. The HAD scores of parents of newly-diagnosed-children group showed that the scores of anxiety and depression were 38.4% and 24.1% higher than the baseline value respectively. The scores of parents’ cultural, recreational orientations and independence increased to normal level in six months after their children received treatment; Fathers’ conflict score decreased to normal level in intervention group (3.0) compared to non-intervention group (4.4). The scores of parents’ anxiety and depression reduced to baseline six months after their children received treatment. We concluded that the psychologic distress of parents was influenced tremendously by their leukemia children. At the beginning of diagnosis their anxiety, depression and conflict were significantly elevated. Most parents could cope with their leukemia children by timely and effective psychological supports. Professional psychological intervention is most helpful in reducing family conflicts.