AIEOP-BFM Acute Lymphoblastic Leukemia Research Articles

ETV6::RUNX1 Acute Lymphoblastic Leukemia: how much therapy is needed for cure?

Recent trials show 5-year survival rates >95% for ETV6::RUNX1 Acute Lymphoblastic Leukemia (ALL). Since treatment has many side effects, an overview of cumulative drug doses and intensities between eight international trials is presented to characterize therapy needed for cure. A meta-analysis was performed as a comprehensive summary of survival outcomes at 5 and 10 years. For drug dose comparison in non-high risk trial arms, risk group distribution was applied to split the trials into two groups: trial group A with ~70% (range: 63.5–75%) of patients in low risk (LR) (CCLSG ALL2004, CoALL 07-03, NOPHO ALL2008, UKALL2003) and trial group B with ~45% (range: 38.7–52.7%) in LR (AIEOP-BFM ALL 2000, ALL-IC BFM ALL 2002, DCOG ALL10, JACLS ALL-02). Meta-analysis did not show evidence of heterogeneity between studies in trial group A LR and medium risk (MR) despite differences in treatment intensity. Statistical heterogeneity was present in trial group B LR and MR. Trials using higher cumulative dose and intensity of asparaginase and pulses of glucocorticoids and vincristine showed better 5-year event-free survival but similar overall survival. Based on similar outcomes between trials despite differences in therapy intensity, future trials should investigate, to what extent de-escalation is feasible for ETV6::RUNX1 ALL.

Pneumocystis jirovecii pneumonia in paediatric acute lymphoblastic leukaemia: A report from the multi-international clinical trial AIEOP-BFM ALL 2009.

Pneumocystis jirovecii can cause life-threatening pneumonia (PjP), and patients with haematological malignancies are at high risk of this infection. Prophylactic measures have significantly decreased morbidity and mortality, but there is a paucity of contemporary data on the incidence and clinical course of PjP in well-defined and homogenous patient populations, such as children suffering from acute lymphoblastic leukaemia (ALL). In the multi-international trial AIEOP-BFM ALL2009, PjP was diagnosed in six children (incidence 1/1000) and was associated with insufficient prophylaxis in five of them. Although none of the patients died of PjP, the long-term impact of the infection is unclear.

CD371-positive pediatric B-cell acute lymphoblastic leukemia: propensity to lineage switch and slow early response to treatment

AbstractIn the effort to improve immunophenotyping and minimal residual disease (MRD) assessment in acute lymphoblastic leukemia (ALL), the international Berlin-Frankfurt-Münster (iBFM) Flow Network introduced the myelomonocytic marker CD371 for a large prospective characterization with a long follow-up. In the present study, we aimed to investigate the clinical and biological features of CD371-positive (CD371pos) pediatric B-cell precursor ALL (BCP-ALL). From June 2014 to February 2017, 1812 pediatric patients with newly diagnosed BCP-ALLs enrolled in trial AIEOP-BFM ALL 2009 were evaluated as part of either a screening (n = 843, Italian centers) or validation cohort (n = 969, other iBFM centers). Laboratory assessment at diagnosis consisted of morphological, immunophenotypic, and genetic analysis. Response assessment relied on morphology, multiparametric flow cytometry (MFC), and polymerase chain reaction (PCR)-MRD. At diagnosis, 160 of 1812 (8.8%) BCP-ALLs were CD371pos. This correlated with older age, lower ETV6::RUNX1 frequency, immunophenotypic immaturity (all P < .001), and strong expression of CD34 and of CD45 (P < .05). During induction therapy, CD371pos BCP-ALLs showed a transient myelomonocytic switch (mm-SW: up to 65.4% of samples at day 15) and an inferior response to chemotherapy (slow early response, P < .001). However, the 5-year event-free survival was 88.3%. Among 420 patients from the validation cohort, 27 of 28 (96.4%) cases positive for DUX4-fusions were CD371pos. In conclusion, in the largest pediatric cohort, CD371 is the most sensitive marker of transient mm-SW, whose recognition is essential for proper MFC MRD assessment. CD371pos is associated to poor early treatment response, although a good outcome can be reached after MRD-based ALL-related therapies.

Impact of Mercaptopurine Metabolites on Disease Outcome in the AIEOP-BFM ALL 2009 Protocol for Acute Lymphoblastic Leukemia.

In the maintenance phase of Associazione Italiana di Ematologia e Oncologia Pediatrica (AIEOP)- Berlin-Frankfurt-Muenster (BFM) acute lymphoblastic leukemia (ALL) 2009 protocol, mercaptopurine (MP) is given at the planned dose of 50 mg/m2 /day; however, dose adjustments are routinely performed to target patients' white blood cells to the optimal range of 2,000-3,000 cells/μL. Pediatric patients with ALL (n = 290, age: median (1st-3rd quartile): 4.8 (3.0-8.1) years; boys: 56.9%) were enrolled mainly in 4 medium-large Italian pediatric hospitals; 14.1% of patients relapsed after a median (1st-3rd quartile) follow-up time of 4.43 (3.82-5.46) years from maintenance beginning. MP metabolites (thionucleotide (TGN) and methyl-derivatives (MMPN)) were measured in the erythrocytes of 387 blood samples of 200 patients by high performance liquid chromatography with ultraviolet detection. Single-nucleotide polymorphisms (SNPs; (rs1800462, rs1800460, and rs1142345 in TPMT gene, rs116855232 in NUDT15, rs1127354, rs7270101, rs6051702 in ITPA, and rs2413739 in PACSIN2) were characterized by Taqman SNP genotyping assays. Cox proportional hazard models did not show an impact of TGN levels and variability on relapse. In contrast, after multivariate analysis, relapse hazard ratio (HR) increased in children with ALL of the intermediate risk arm compared with those in standard risk arm (3.44, 95% confidence interval (CI), 1.31-9.05, P = 0.012), and in carriers of the PACSIN2 rs2413739 T allele compared with those with the CC genotype (heterozygotes CT: HR, 2.32, 95% CI, 0.90-5.97, P = 0.081; and homozygous TT: HR, 4.14, 95% CI, 1.54-11.11, P = 0.005). Future studies are needed to confirm the lack of impact of TGN levels and variability on relapse in the AIEOP-BFM ALL trials, and to clarify the mechanism of PACSIN2 rs2413739 on outcome.

Hypersensitivity Reactions to Native E. coli L-asparaginase in Children With Acute Lymphoblastic Leukemia Treated in Trial ALL-BFM 2000: Impact of Treatment Schedule and Type of Glucocorticoid in Induction.

Hypersensitivity Reactions to Native E. coli L-asparaginase in Children With Acute Lymphoblastic Leukemia Treated in Trial ALL-BFM 2000: Impact of Treatment Schedule and Type of Glucocorticoid in Induction.

Effect of two additional doses of intrathecal methotrexate during induction therapy on serious infectious toxicity in pediatric patients with acute lymphoblastic leukemia.

Although initial central nervous system (CNS) involvement is rarely detected in childhood acute lymphoblastic leukemia (ALL), risk-adapted CNS-directed therapy is essential for all patients. Treatment intensity depends on the initial CNS status. In the AIEOP-BFM ALL 2009 trial, patients with cytomorphologic detection of leukemic blasts in initial cerebrospinal fluid were classified as CNS2 or CNS3 and received five intrathecal doses of methotrexate (MTX) in induction therapy compared to patients with CNS1 status (no blasts detected) who received three doses. The impact of additional intrathecal (IT) MTX on systemic toxicity in induction therapy is unknown. Between June 1st 2010 and February 28th 2017, a total of 6,136 ALL patients aged 1-17 years were enrolled onto the AIEOP-BFM ALL 2009 trial. The effect of three versus five doses of IT MTX during induction therapy on the incidence of severe infectious complications was analyzed. Among 4,706 patients treated with three IT MTX doses, 77 (1.6%) had a life-threatening infection during induction as compared to 59 of 1,350 (4.4%) patients treated with five doses (P<0.001; Odds Ratio 2.86 [95% Confidence Interval 1.99-4.13]). In a multivariate regression model, treatment with additional IT MTX proved to be the strongest risk factor for life-threatening infections (Odds Ratio 2.85 [1.96-4.14]). Fatal infections occurred in 16 (0.3%) and 38 (1.6%) patients treated with three or five IT MTX doses, respectively (P<0.001). As the relevance of additional intrathecal MTX in induction for relapse prevention in CNS2 patients is unclear, doses of intrathecal therapy have been reduced for these patients. (Clinicaltrials.gov identifiers: NCT01117441 and NCT00613457).

Chemotherapy-related hyperbilirubinemia in pediatric acute lymphoblastic leukemia: a genome-wide association study from the AIEOP-BFM ALL study group

BackgroundCharacterization of clinical phenotypes in context with tumor and host genomic information can aid in the development of more effective and less toxic risk-adapted and targeted treatment strategies. To analyze the impact of therapy-related hyperbilirubinemia on treatment outcome and to identify contributing genetic risk factors of this well-recognized adverse effect we evaluated serum bilirubin levels in 1547 pediatric patients with acute lymphoblastic leukemia (ALL) and conducted a genome-wide association study (GWAS).Patients and methodsPatients were treated in multicenter trial AIEOP-BFM ALL 2000 for pediatric ALL. Bilirubin toxicity was graded 0 to 4 according to the Common Toxicity Criteria (CTC) of the National Cancer Institute. In the GWAS discovery cohort, including 650 of the 1547 individuals, genotype frequencies of 745,895 single nucleotide variants were compared between 435 patients with hyperbilirubinemia (CTC grades 1-4) during induction/consolidation treatment and 215 patients without it (grade 0). Replication analyses included 224 patients from the same trial.ResultsCompared to patients with no (grade 0) or moderate hyperbilirubinemia (grades 1-2) during induction/consolidation, patients with grades 3-4 had a poorer 5-year event free survival (76.6 ± 3% versus 87.7 ± 1% for grades 1-2, P = 0.003; 85.2 ± 2% for grade 0, P < 0.001) and a higher cumulative incidence of relapse (15.6 ± 3% versus 9.0 ± 1% for grades 1-2, P = 0.08; 11.1 ± 1% for grade 0, P = 0.007). GWAS identified a strong association of the rs6744284 variant T allele in the UGT1A gene cluster with risk of hyperbilirubinemia (allelic odds ratio (OR) = 2.1, P = 7 × 10− 8). TT-homozygotes had a 6.5-fold increased risk of hyperbilirubinemia (grades 1-4; 95% confidence interval (CI) = 2.9-14.6, P = 7 × 10− 6) and a 16.4-fold higher risk of grade 3-4 hyperbilirubinemia (95% CI 6.1-43.8, P = 2 × 10− 8). Replication analyses confirmed these associations with joint analysis yielding genome-wide significance (allelic OR = 2.1, P = 6 × 10− 11; 95% CI 1.7-2.7). Moreover, rs6744284 genotypes were strongly linked to the Gilbert’s syndrome-associated UGT1A1*28/*37 allele (r2 = 0.70), providing functional support for study findings. Of clinical importance, the rs6744284 TT genotype counterbalanced the adverse prognostic impact of high hyperbilirubinemia on therapy outcome.ConclusionsChemotherapy-related hyperbilirubinemia is a prognostic factor for treatment outcome in pediatric ALL and genetic variation in UGT1A aids in predicting the clinical impact of hyperbilirubinemia.Trial registrationhttp://www.clinicaltrials.gov; #NCT00430118.

Invasive fungal diseases impact on outcome of childhood ALL – an analysis of the international trial AIEOP-BFM ALL 2009

In children with acute lymphoblastic leukemia (ALL), risk groups for invasive fungal disease (IFD) with need for antifungal prophylaxis are not well characterized, and with the advent of new antifungal compounds, current data on outcome are scarce. Prospectively captured serious adverse event reports of children enrolled in the international, multi-center clinical trial AIEOP-BFM ALL2009 were screened for proven/probable IFD, defined according to the updated EORTC/MSG consensus definitions. In a total of 6136 children (median age 5.2 years), 224 proven/probable IFDs (65 yeast and 159 mold) were reported. By logistic regression, the risk for proven/probable IFDs was significantly increased in children ≥12 years and those with a blast count ≥10% in the bone marrow on day 15 (P < 0.0001 each). Proven/probable IFDs had a 6-week and 12-week mortality of 10.7% and 11.2%, respectively. In the multivariate analysis, the hazard ratio for event-free and overall survival was significantly increased for proven/probable IFD, age ≥12 years, and insufficient response to therapy (P < 0.001, each). Our data define older children with ALL and those with insufficient treatment-response at high risk for IFD. As we show that IFD is an independent risk factor for event-free and overall survival, these patients may benefit from targeted antifungal prophylaxis.

Impact of Additional Intensive L-Asparaginase Therapy during Consolidation Phase for High-Risk Acute Lymphoblastic Leukemia: Results of a Randomized Controlled Trial in the AIEOP-BFM ALL 2009 Protocol

Introduction Treatment of childhood Acute Lymphoblastic Leukemia (ALL) with unfavorable features remains a major challenge. An earlier intervention, aiming at lowering Minimal Residual Disease (MRD) levels, might reduce the risk of relapse and the indication to allogeneic hematopoietic stem cell transplantation (HSCT) in 1st complete remission (CR). The AIEOP-BFM ALL 2009 protocol included a randomized study for patients with high-risk (HR) features, to investigate if an intensive and continuous exposure to pegylated L-Asparaginase (Peg-L-ASP), given on top of BFM consolidation phase IB, could decrease the levels of MRD and to influence long term outcome. Patients and methods From June 1, 2010 to February 28, 2017, 6136 patients with Ph’ negative ALL, aged between 1-17 years, were enrolled in the AIEOP-BFM ALL 2009 Study (Eudract number 2007-004270-43). Participating study centers were in Australia, Austria, Czech Republic, Germany, Israel, Italy and Switzerland.1097 were classified as HR at the end of induction phase IA (day 33), on the basis of evidence ofKMT2A-AFF1 rearrangement, hypodiploidy (<45 chromosomes, or DNA-index <0.8), prednisone poor response (PPR), poor bone marrow (BM) response at day 15 (i.e. ≥10% blasts by MFC), failure to achieve morphological CR. Treatment consisted of a 7-day prephase with prednisone and 1 intrathecal dose of methotrexate (ITMTX), an induction protocol IA with prednisone or dexamethasone, vincristine, daunomycin, Peg-L-ASP (2 doses), ITMTX and, only in PPR T-ALL cyclophosphamide. Patients eligible were randomized to receive or not 4 weekly doses of Peg-L-ASP (2500 IU/sqm x 4) on top of consolidation phase IB, which included 6-MP, cyclophosphamide, cytosine arabinoside and ITMTX. Thereafter, patients received 3 blocks of non-cross-resistant drugs, 3 reinduction phases with or without cranial radiotherapy (12 Gys), and maintenance therapy till 24 months from diagnosis. The primary endpoint was PCR-MRD reduction to less than 5x10-4at the end of phase IB; secondary endpoint was Event-Free Survival (EFS). Results Overall, 1097 patients were at HR at the end of induction phase IA and 809 (79.2%) were randomized to receive (404) or not (405) 4 doses of Peg-L-Asp during consolidation phase IB. The primary endpoint could be assessed in 732 patients (90.5%). By ITT analysis, the proportion of patients with PCR MRD ≥ 5x10-4 at the end of consolidation phase IB in the experimental arm (EA) vs the control arm (CA) was 13.9% vs 17.0% with no significant difference (p-value=0.25).There were more toxic deaths during consolidation phase IB among patients randomized to Peg-L-Asp (n=14, 12 of them due to infections), vs controls (n=1): 7 of them, all in the EA, however, occurred before starting the randomized treatment. Deaths in CR after phase IB were 24 vs 24 in the EA vs CA (of whom 10 vs 12 were after HSCT in CR1, performed in 87 and 90 patients respectively). In the EA and CA the overall 5-year EFS by ITT was 70.4% (2.3) and 75.0% (2.2) (p-value=0.18), the 5-year cumulative incidence of death in CR was 9.5% (1.5) vs 5.7% (1.2) (p-value=0.08) and that of relapse was 17.7% (1.9) vs 17.2% (1.9), respectively (p-value=0.94). Non HSCT related deaths were due to infections in 23/28 in the EA, and in 10/13 in CA patients. The 5-year overall survival probability was 81.5% (2.0) and 84.0% (1.9) respectively (p-value=0.25). Discussion In this randomized study, results (ITT analysis) show a very modest, not significant decrease in the percentage of patients with high MRD in the EA, and no significant difference in EFS between the two arms. The trial results suggest that the additional administration of intensive Peg-L-Asp in phase IB is associated with higher, although not statistically significant, mortality in CR and very similar relapse incidence. However, some fatal events in phase IB occurred very early after randomization, before the start of the planned treatment with Peg-L-Asp. Importantly, in both analyses, by ITT and by treatment given, there is no evidence of benefit on reduction of relapses by the additional therapy with Peg-L-ASP in phase IB. These findings indicate that an early intensification of chemotherapy with Peg-L-Asp in patients defined as HR does not improve the outcome of children with HR ALL treated according to AIEOP-BFM regimen, thus suggesting that further improvements with conventional chemotherapy might be difficult in the context of intensive treatment protocols. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

First Description of a Frameshift PAX5 Germline Variant in Two Siblings with B-Cell Precursor Acute Lymphoblastic Leukemia

Introduction: Recent studies have highlighted that about 5% of children with Acute Lymphoblastic Leukemia (ALL) carry a germline mutation that predispose to leukemogenesis. PAX5 somatic mutations are frequently identified on sporadic B-ALL, but recently pathogenic PAX5 germline variants have been described in families with recurrence of B-ALL (Shah et al. 2013; Auer et al. 2014; Yazdanparast et al. 2015, Duployez et al. 2021). We describe the case of two siblings both affected by B-ALL with evidence of a new familial PAX5 variant. Case A was a male, 13 years old, diagnosed with B-II ALL. He was enrolled into the AIEOP-BFM ALL 2009 treatment protocol and according to minimal residual disease (MRD) levels he was stratified as medium risk. Eight years later, his sister (14yrs, named as case B) was diagnosed with B-II ALL, and stratified in the medium risk group of AIEOP-BFM ALL 2017 protocol. The two patients were otherwise healthy and borne from healthy unrelated parents; no history of hematological malignancies was reported across the pedigree. Methods: We performed a custom Next Generation Sequencing ALL predisposition panel on DNA extracted from bone marrow samples of both disease onset and remission phase (defined as negativity by PCR-MRD, 10-4 sensitivity). The libraries were performed following Nextera Flex for Enrichment and sequenced on Illumina Nextseq550 platform; the analyses were carried out by Congenica software. The variants of interest were validated and investigated in the family pedigree through PCR and Sanger sequencing. Digital-MLPA testing 55 ALL hot spot genes (including all PAX5 exons) was performed. Results: Through the NGS analysis, we showed that both patients shared a novel germline heterozygous PAX5 variant (c.548delG, p.Gly183AlafsTer84); moreover, the disease samples of both patients carried an additional somatic PAX5 variant (c.239C>G, p.Pro80Arg; variant fraction: 29% in Case A and 38% in Case B). The Digital-MLPA using the ALL panel, excluded the presence of shared copy number alterations (CNV) and, specifically, did not identified CNV alterations involving the PAX5 locus. The familiar segregation showed a paternal origin of the germline PAX5 variant (p.Gly183AlafsTer84); the mother was found to be wild-type. The PAX5 variant was also found in the DNA sample of the patients' grandmother of the paternal branch; a healthy brother of probands (24 years old) was not tested. To investigate a possible impairment of PAX5 function in the development of the B-cell lineage, we evaluated in both parents the B-cell repertoire on peripheral blood fresh mononuclear cells by FACS analysis. Interestingly, the father has a significant reduction on mature IgD+/CD27+ B-cells with a concomitant accumulation of IgD+/CD27- naïve B-cells. Conclusions: We are reporting a novel germline PAX5 p.Gly183AlafsTer84 in a family with recurrence of B-ALL. In contrast to the previous PAX5 described cases that carry an inherited missense variant resulting in a modest attenuation of PAX5 activity, our patients present a germline frameshift variant that brings to premature stop of the protein, suggesting a more destructive impact on PAX5 activity. As in the other reported cases, our patients presented an additional somatic event on PAX5 that further compromise PAX5 function, leading to the establishment of the leukemic clone. Interestingly and different from the literature, the two siblings share the same PAX5 p.Pro80Arg somatic variant, that has been characterized by Gu as a pathogenic variant impairing B lymphoid development and promoting leukemogenesis (Gu et al. 2019). The detailed molecular characterization, including Whole Transcriptome NGS analysis, is ongoing to investigate whether a PAX5-specific signature can be identified. Moreover, a single-cell NGS approach on remission samples of the two siblings could help to identify specific important pathways and altered gene suppression profiles, preceding full leukemia. Consent-dependent characterization of additional family members are ongoing to assess the penetrance of leukemia associated to this new variant and possible additional features. The identification and characterization of germline PAX5 variants is of high interest in the context of leukemia predisposition because of its clinical implications in terms of familial genetic counselling and because of its contribution in the understanding ALL pathogenesis.

Hypersensitivity Reactions to Native E. coli L-Asparaginase in Children with Acute Lymphoblastic Leukemia May Vary By Treatment Schedule and Type of Glucocorticoid in Induction: Results of Trial ALL-BFM 2000

Background: Asparaginase (ASNase) has become a key component of treatment for acute lymphoblastic leukemia (ALL). The ability of this agent to eliminate leukemic blasts by utilizing the special feature that such cells are largely unable to synthesize the otherwise non-essential amino acid asparagine means that blasts cannot survive in an environment in which this amino acid has been depleted. Currently, there are three ASNase products commercially available sharing the same mechanism of action. The oldest one is the purified native enzyme extracted from E. coli. This product was further developed and refined by conjugation with polyethylene glycol (PEG-L-Asparaginase, PEG-ASNase) to prolong half-life and to reduce immunogenicity. A third ASNase product derived from the bacterium Erwinia chrysanthemi (Erwinase) is structurally different from the E. coli ASNase products. The main reason for limitation in the use of ASNase is the rather frequent occurrence of hypersensitivity reactions (HSR). Such reactions require the replacement of the ASNase product used with alternative preparations. The incidence of clinical HSR to ASNase varies with the type of ASNase product, the treatment schedule (number of ASNase doses, extended ASNase-free intervals) and the administration route (intravenously [IV] or intramuscularly [IM]). This retrospective study was primarily performed to demonstrate the dynamics and the interdependence when using ASNase in a complex multiagent treatment protocol. It aimed to evaluate the incidence of clinically apparent HSR in the ALL-BFM 2000 trial which was part of the cooperative trial AIEOP-BFM ALL 2000. Methods: Clinical hypersensitivity reactions (HSR) to native E. coli ASNase were retrospectively analyzed in a cohort of children with acute lymphoblastic leukemia (ALL) focusing on the impact of concurrent chemotherapy schedules in the risk-adapted and randomized treatment protocol and evaluating the prognostic relevance of asparaginase discontinuation. Patients: Patients with ALL aged 1-18 years were treated risk-stratified with standard-risk (SR), medium-risk (MR) and high-risk (HR) regimens in trial ALL-BFM 2000. Schedule of intravenously administered native E. coli ASNase varied by risk group and randomization arm. Randomizations compared the glucocorticoids prednisolone and dexamethasone in induction and different reintensification regimens (Möricke A et al, BLOOD 127(17): 2101-2112 (2016)). In this trial, native E. coli ASNase was administered IV as first-line product during the induction and reinduction phases. In the high-risk arm, the phase of intensified consolidation included additional E. coli ASNase doses. PEG-ASNase (IV) or Erwinase (IM) were used as second- or third-line products after HSR occurring with native E. coli ASNase or the respective second-line product. Results: HSR occurred in 40% (492/1207) of all evaluable patients. One-year cumulative incidence (CI-HSR±standard error) was 42.3±1.5% (40.6±2.5% in SR, 37.0±1.9% in MR, 65.2±3.8% in HR). Dexamethasone-treated SR and MR patients had significantly higher CI-HSR (48.5±4.3% and 45.9±3.2%) than prednisolone-treated patients (32.3±3.7% [p=0.007] and 31.2±2.9% [p<0.001]); no such difference was seen among HR patients who underwent multiple exposures to the drug (Fig. 1). Post-induction randomizations showed that one week of dexamethasone before asparaginase re-exposition in reinduction did not protect from clinical HSR in the SR/MR arm. Randomization in HR revealed an effect of the type of concurrent chemotherapeutic drugs on the risk of HSR. Discontinuation of ASNase occurred in 6.1% of all patients. No disadvantage in leukemia outcome could be detected for these patients. Conclusion: The results in our study demonstrate the complex interdependencies contributing to the risk of HSR to E. coli asparaginase in a multiagent chemotherapy protocol. The differential effects of dexamethasone and prednisolone in induction generated new hypotheses about the role of glucocorticoids in sensitization or achievement of immune tolerance to asparaginase. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

P362: MINIMAL-LOSS-CYTOMETRY FOR EVALUATION OF CEREBROSPINAL FLUID IN PEDIATRIC ACUTE LYMPHOBLASTIC LEUKEMIA. SINGLE CENTER STUDY.

Background: Central nervous system (CNS) involvement at diagnosis of acute lymphoblastic leukemia (ALL) represents a risk factor for CNS relapse. Currently, examination of the cerebrospinal fluid (CSF) is performed and evaluated quantitatively using a counting chamber and qualitatively using cytomorphological analysis of cytospin. Recently, flow cytometry (FC) evaluation of CSF was shown to provide more reliable results and to predict relapse risk (de Haas et al., 2021, Thastrup et al., 2020). Independently of these studies, we applied a lyse-no wash technique aiming at minimizing cell loss during preparation and at improved cell concentration estimate. Aims: Our aim was to compare our findings with morphological criteria and published data and correlate them with clinical outcome in pediatric patients with ALL. Methods: We included pediatric patients with newly diagnosed ALL (04/2014 – 01/2022), whose CSF examination was performed in CLIP laboratory (n=208). Patients were classified as B cell precursor ALL (BCP ALL; 85%) and T ALL (15%) and treated according to following protocols: AIEOP-BFM ALL 2009 (n=118), AIEOP-BFM ALL 2017 (n=71), EsPhALL(n=8), Interfant06 (n=5), off-protocol (n=6). During follow up, 14 patients relapsed (5 CNS incl. combined, 9 non-CNS). CSF (drawn into tube with no fixative agent and processed within 2 hours) was concentrated by centrifugation and incubated with combination of antibodies (CD20/CD10/CD45/CD34/CD19/CD3/Syto41 in BCP ALL and CD4/CD99/CD5/CD3/CD7/CD16 + 56/CD8/CD45/Syto41 in T ALL). Residual erythrocytes were lysed with NH4Cl solution, followed by immediate acquisition on BD FACS Lyric or BD LSRII cytometers. Blasts were quantified to initial CSF volume. Cluster of 20 events was required to assign samples as FC positive (FC+). In parallel, CNS status was concluded according to morphological criteria, being classified as CNS1 (no blasts), CNS2a/b (≤5 WBC/μL with blasts), CNS2c/CNS3 (≥5 WBC/μL with blasts) according to treatment protocol guidelines. Continuous variables were compared with the Mann–Whitney test, survival data were analyzed using Mantel-Cox test. P values<0.05 were considered as significant. Results: Using FC, we identified atypical blasts in 50 out of the 208 analyzed samples (24%). Median of analyzed CSF volume was 367µL (range 143-1033µL), which enabled median sensitivity 0.054 events/µL (range 0.02-0.14ev/µL). Mean blast concentration was 0.47ev/µL (range 0-25ev/µL) in all samples and 2 ev/µL (range 0.01-25ev/µL, median 0.53ev/µL) in FC+ samples. We observed significantly higher blast concentration in patients with T ALL, in patients with CNS2 and CNS3 and those who subsequently relapsed. Patients with FC+ had significantly lower relapse-free survival than FC- patients (55% vs 94%), which provided better separation than CNS status (68% vs.85% for CNS2/3 vs. CNS1, respectively). Image:Summary/Conclusion: Using fresh CSF sample with lyse-no wash protocol enabled us to detect approximately 10x higher blast count (0.24ev/µL in FC+CNS1 and 1.25ev/µL in FC+CNS2/3) than was recently published (0.015ev/µL in FC+CNS1 and 0.073ev/µL in FC+CNS2/3 in Thastrup et al., 2020), and thus it better correlates to cytomorphological data. Interestingly, the proportion of FC+ patients in our cohort is comparable to (24% vs 25%), showing that although in our method we lose fewer cells, we identify the same proportion of FC+ patients. The presented study shows a higher impact of FC positivity on relapse risk than described previously. The disadvantage of our method is a need for a fresh sample, which is investigated locally. Supported by UNCE/MED/015, NU20J-07-00028.

PB1760: AZACITIDINE AS TREATMENT OF KMT2A INFANT ACUTE LYMPHOBLASTIC LEUKEMIA: SINGLE CENTER EXPERIENCE

Background: KMT2A rearranged (r) infant acute lymphoblastic leukemia (ALL) is characterized by a very poor prognosis, particularly after relapse. Intensive chemotherapy does not improve outcome and novel therapies are under investigation for this dismal disease. Recent studies reported an hypermethylated DNA profile in KMT2Ar ALL, leading to repression and silencing of tumor suppressor genes. Hypomethylating agents (HMA) have been hypothesized to exert an antileukemic activity restoring normal gene expression. Aims: Here we report the off-label administration of Azacitidine (AZA) in two little patients aged 1 month old and 26 days old respectively, diagnosed with KMT2Ar infant ALL. Methods: Both patients were treated according to the high risk arm of the AIEOP-BFM ALL 2017 protocol as front-line therapy. AZA was administered every 21 days (d) for 4 cycles (cy), with the following schedule: 55 mg/m2 for 5 d, 55 mg/m2 for 7 d, 75 mg/m2 for 5 d, 75 mg/m2 for 7 d, respectively; prednisone 1 mg/m2 was associated to each cycle. Results: Patient 1 was diagnosed with infant B ALL t(11;19)/KMT2A-MLLT1r, high-risk (HR) for age, hyperleucocytosis and KMT2Ar. At the end of induction (EOI) he was in hematologic remission with molecular minimal residual disease (MRD) and t(11;19) still detectable. The child received consolidation chemotherapy followed by 2 cy of bispecific CD3-CD19 antibody (ab) Blinatumomab and underwent allogeneic hematopoietic stem cell transplant (HSCT) from matched unrelated donor (MUD) with detectable MRD. Two months after transplant patient relapsed with 68% CD19 CD22 negative blasts in bone marrow. Due to early relapse and no target therapy available, AZA was administered. Grade 4 cytopenia and pneumonia were main adverse events. Patient achieved MRD complete remission (CR) and underwent to haploidentical HSCT from mother, followed by 3 courses of 40 mg/m2 AZA maintenance for 5 d, every 21 d. Patient is alive, in MRD negative CR3, 3 months after SCT. Patient 2 was diagnosed with infant B ALL t(4;11)/KMT2A-AFF1, HR for age, hyperleucocytosis and KMT2Ar. At the EOI bone marrow showed blasts 25% detected by flow cytometry. Patient received one courses of chemotherapy according to the HR protocol, followed by 2 cy of Blinatumomab. The child achieved MRD negative CR and underwent HSCT from MUD. Thirty two days after HSCT, patient relapsed with 60% BM blasts detected by flow cytometry. Due to early relapse after HSCT, he received 2 cy of anti-CD22 ab Inotuzumab, achieving CR without platelet recovery (CRi). Nevertheless, t(4;11) was positive again at the end of the 2nd cy. Patient then received AZA as 3th-line therapy achieving CRi after 2nd cy. However, due to severe cytopenia and gastrointestinal toxicity, patient received 3 out of 5 days treatment of the 3th cy and the 4th cy was delayed. Patient relapsed after the 4thcourse. Summary/Conclusion: In our experience, single agent AZA alone was effective and well tolerated in heavily pretreated patients. To our knowledge, single agent HMA in KMT2Ar ALL has been reported only in one patient. Preclinical studies have revealed heterogeneous results when HMA are combined with cytotoxic drugs; results of two pilot trials of AZA combined with chemotherapy are still ongoing. Further clinical trials could clarify the optimal schedule, timings and dosage of HMA.

Conventional and Flowcytometric Minimal Residual Disease Based Risk Stratified Post Remission Therapy Improves Survival in Ph-Negative Pediatric Acute Lymphoblastic Leukemia- Single Centre Experience from South India

Background: Children with Acute Lymphoblastic Leukemia (ALL) in Low Middle income countries(LMIC) face major challenges including treatment abandonment and poor overall survival (OS) and event-free survival (EFS).The preliminary data and results of outcome of Pediatric ALL from our pediatric hematology-oncology center which follows BFM based ALLtreatment protocol has been published in 2015 in ASH forum.Due to non availability of Polymerase chain reaction (PCR)based Minimal residual disease(MRD)analysis, we use multiparametric flowcytometry (FCM) based MRD analysis for remission assessment and risk stratification in our patients. Within resource constraints, we present evidence that outcomes comparable with that seen in high income (HIC)and upper middle income countries(UMIC) can be achieved with a post remission therapy guided by a risk stratification incorporating FCM MRD.Methods: Following IRB approval,an ambidirectional cohort study was performed using clinical information and outcomes of all patients aged 1to 14 years treated for newly diagnosed B- or T-ALL between January 1,2015 and December 31, 2020. AIEOP BFM ALL 2009 protocol with modifications was followed as the institutional protocol in all patients.The treatment algorithm is mentioned in Figure 1. Patients who underwent multiparametric FCM MRD analysis at the end of Induction IA and whose 6 months follow up details were available were included in the analysis. Patients with Ph positive ALL and those who died during Induction IA were excluded. FCM MRD analysis was performed after Induction IA on Day 33.MRD level above 0.01% was considered positive.MRD assessment was repeated following Induction IB on Day 74 in patients who had MRD positivity on Day 33. At the end of Induction IB patients were risk stratified into High risk(HR) and non High risk(Non HR).HR features included Hypodiploidy(<45chromosomes), Positivity for MLL/AF4 or t(4:11),poor prednisolone response(absolute blast count in peripheral blood ≥1 x 10^9/l on Day 8 of initiation of prednisolone) and non remission on Day33. Patients who had persistent MRD positive on Day74 were reallocated to high risk. Treatment with HR protocol was initiated for high risk patients whenever financially manageable.Statistical analysis was done using SPSS version 21 OS and EFS were assessed by Kaplan-Meier method Patients were censored at last follow-up.Results: Median follow-up time was 33.5(4-102) months . Study had 102(n=102) consecutive Ph negative patients who underwent induction therapy. Six patients (5.88%) died during induction IA;96(n=96) children who continued treatment were included in further analysis. The mean age at diagnosis was 6(1-14) years .Forty seven (48.95%)patients were male..B-ALL n=84(87.5%) and T ALL n=12.(12.5%). Four patients(4.16%) had CNS disease at diagnosis. Three children (3.13%) had high risk cytogenetics.Ten children (10.42%)in the cohort had poor prednisolone response. Five patients(5.20%) didnot achieve morphological remission on Day33. Fifteen (15.63%) patients were risk stratified as HR during IA. Four more (4.16%) were reallocated to HR in view of persistent MRD positivity after Induction IB. At median follow-up, the OS was 95.35%±2.65(95% CI 90.16-100.54) and the EFS 94.48%±2.74 (95% CI, 89.11%-99.84%).Female gender predicted better EFS (p=0.042).The EFS of patients without CNS disease at presentation was significantly better(93.5% Vs 23.2% p=0.000). The EFS at median follow up of patients in re HR cohort was 64.19% Vs 97.81% in non HR (p=0.010). EFS by risk stratification is shown in Figure II.Conclusion:Our study suggests that FCM MRD can be successfully incorporated into the treatment algorithm in a resource limited setting. With FCM MRD were able to identify an additional subset of HR patients who otherwise would have been stratified into non HR group. In a prospective cohort , FCM MRD could be tested at an earlier time point in IA induction to facilitate identification of early drug responsive patients with lowest risk of relapse. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

NGS-Based MRD Quantitation: An Alternative to qPCR Validated on a Large Consecutive Cohort of Children with ALL

Together with multicolor flow cytometry, quantitation of clonal immunoglobulin (IG) and T-cell receptor (TR) gene rearrangements represents the current standard for the detection of minimal / measurable residual disease (MRD) in treatment protocols for pediatric acute lymphoblastic leukemia (ALL) patients. Despite the adoption of next generation sequencing (NGS) in the routine identification of clonal IG/TR gene rearrangements as markers for MRD detection, real-time quantitative (q)PCR is still the standard for MRD quantitation in follow-up samples. So far, no large-scale direct comparison of qPCR- and NGS-based MRD quantitation has been performed.We compared qPCR- and NGS-MRD evaluation in a cohort of children with B-cell precursor (BCP) ALL treated on the AIEOP-BFM ALL 2009 protocol and assessed the feasibility and relevance of this method for the stratification at day 33 (EOI).In total, 459 patients were diagnosed with BCP-ALL from 2010 to 2018, and 437 of them were included in our study based on the availability of residual DNA material isolated from day 33 bone marrow aspirates and having at least one IG/TR MRD marker detectable by standard qPCR with protocol-required sensitivity of 10 -4. Sequencing libraries were prepared according to the EuroClonality-NGS group SOP (Brüggemann et al, Leukemia 2019) with the total DNA input normalized to the equivalent of 150,000 nucleated cells to reach MRD sensitivity of 10 -5 and sequenced on Illumina NovaSeq and MiSeq instruments. In total of 780 IG/TR markers evaluated by both NGS and qPCR. Sequencing data were analyzed using the ARResT/Interrogate (Bystry et at, Bioinformatics 2017) pipeline and a custom bioinformatic analysis process and the NGS-MRD results were normalized to the EuroClonality-NGS central in-tube quality/quantification control (cIT-QC; Knecht et al, Leukemia 2019).From the total 780 IG/TR MRD markers evaluated by both methods, 629 (80.6%) were concordant with 242 markers being MRD positive and 387 negative. From 82 markers that were only positive by qPCR and not by NGS, 76 were positive below the quantitative range (positive non-quantifiable). Specificity analysis was performed for each marker by searching for the junction sequence across the dataset of all patients' NGS results. Based on these results, 22 out of 82 markers positive only by qPCR were classified as potentially unspecific (false positive) and similarly 32 unspecific markers were identified among the 69 positive only by NGS. This was also supported by unspecific amplification of the polyclonal control in 27 out of these 32 corresponding qPCR systems, in some cases leading to qPCR negative classification determined by the EuroMRD guidelines.Overall stratification of patients based only on day 33 MRD by qPCR or NGS was concordant in 76% of patients by both methods, while in 19% of patients, NGS-MRD quantitation led to the assignment to a lower-risk group, mainly due to the elimination of false-positive results. Furthermore, analysis of all positive markers across all patients' NGS libraries showed, that one out of 10 markers (mainly in the IGK, TRG and TRD loci) used for qPCR-MRD stratification did not provide satisfactory specificity, although they fully met EuroMRD criteria during the optimization of qPCR patient-specific assays.Our results show that NGS-MRD is highly concordant with traditional qPCR-based strategy and has comparable sensitivity and clinical value in the setting of a BFM-based clinical protocol, while being less laborious and providing significantly more specific results and additional information on the IG/TR repertoire (Kotrova et al, Blood 2016). Our study also emphasizes the importance of selecting MRD markers of adequate specificity at diagnosis. Currently, this selection can be assisted by these broad sequencing data on IG/TR repertoire of large number of patients. Based on these results, we propose that frontline NGS-MRD evaluation developed by the EuroClonality-NGS working group can be used as an alternative to traditional qPCR-based MRD quantitation in future MRD-based treatment protocols.Supported by grants NU20-03-00284 and NU20-07-00322 from the Czech Health Research Council and 534120 from Charles University. All methods were established through collaboration within the EuroClonality-NGS and EuroMRD groups. Disclosuresvan der Velden: Agilent: Research Funding; Navigate: Other: Service Level Agreement; Janssen: Other: Service Level Agreement; EuroFlow: Other: Service Level Agreement, Patents & Royalties: for network, not personally; BD Biosciences: Other: Service Level Agreement. Brüggemann: Incyte: Other: Advisory Board; Janssen: Speakers Bureau; Amgen: Other: Advisory Board, Travel support, Research Funding, Speakers Bureau. Langerak: Erasmus MS, University Medical Center: Current Employment; F. Hoffmann-La Roche Ltd/Genentech, Inc.: Research Funding; Gilead: Research Funding; Janssen: Speakers Bureau.

Genome-Wide Association Study (GWAS) of High-Risk Minimal Residual Disease and Relapse in B-Cell Acute Lymphoblastic Leukemia on Trial AIEOP-BFM ALL 2000

▪A significant proportion of patients with pediatric acute lymphoblastic leukemia (ALL) still suffers from relapse and therapy-related toxicities. Thus, the focus of further improvement of therapy for them is not only cure, but minimizing both short-term and long-term therapy-associated toxicity. This can be achieved by better personalized adjustment of therapy to the risk of relapse. Continuing research has identified numerous features with prognostic potential, including clinical and genetic variables as well as early response to chemotherapy being associated with treatment outcome. While most research activities on genetic prognostic factors have focused on somatic features, relatively little is known on hereditary contributions to treatment response and outcome in pediatric ALL. Therefore, we conducted a genome-wide association study (GWAS) of high-risk minimal residual disease (MRD) and relapse in B cell precursor (BCP) ALL patients treated on trial AIEOP-BFM ALL 2000.On this trial, patients were stratified into three risk groups, mainly by MRD analyses. MRD standard-risk (MRD-SR) patients were MRD-negative on treatment days 33 and 78, MRD high-risk (MRD-HR) patients had residual disease of ≥5x10-4 on treatment day 78. All the remaining MRD results were considered MRD intermediate-risk (MRD-IR). Besides MRD, established high-risk parameters were also retained for stratification: prednisone poor-response, induction failure, positivity for t(4;11)/ MLL-AF4 . Treatment used standard drugs and, in some of the patients, cranial irradiation and/or hematopoietic stem cell transplantation. DNA isolated from remission samples was genotyped using Illumina Human1M-Duo BeadChips containing 1.048.711 single SNV markers. For GWAS analyses gPLINK v2.050, PLINK v.1.09 and RStatistics v3.3.1 were used. The analysis on MRD was conducted in 69 MRD-HR patients (cases) and in 1034 patients of the MRD-SR and MRD-IR groups (controls). The analysis on relapse employed 145 relapsed (cases) and 1007 non-relapsed patients (controls). After quality control assessment, the final analyses included 749.262 SNPs among 62 MRD-HR patients compared to 927 controls and 129 relapsed patients compared to 925 controls.In the analysis on MRD-HR five SNVs reached genome-wide significance at P ≤5×10−8. An intronic SNV (rs1570765) in STARD13 demonstrated the strongest association (P =1.73x10-10; odds ratio, OR=5.06). Two additional SNVs with P ≤5×10−8 in STARD13 were in linkage disequilibrium with rs1570765. The two remaining SNVs with P ≤5×10−8 were located in FEZ2 and NRIP1. Five additional SNVs demonstrated significant levels of ≤1×10−6 and were associatedwith the following genes: STARD13 , VIT and FICD . STARD13 is located on chromosome 13, belongs to the family of RhoGap proteins and is frequently lost in hepatocellular and breast carcinomas. Its RhoGAP domain activates members of the Ras superfamily which leads to a suppression of cell growth and differentiation (Lin et al. Oncogene 2010). In the GWAS approach on relapse three SNVs reached significant levels of ≤1×10−6 and were associatedwith the genes ASB3 (rs1549749, P =3.51x10-7; rs12465316, P =4.26x10-7) on chromosome 2 and NDN (rs9972595, P =1.04x10-6) on chromosome 15. Twenty additional SNVs demonstrated significance levels of ≤1×10−5. ASB3 codes for a member of the ankyrin repeat and SOCS box-containing (ASB) family of proteins containing ankyrin repeat sequence and a SOCS box domain. The latter serves to couple suppressor of cytokine signalling (SOCS) proteins and their binding partners with the elongin B and C complex, possibly targeting them for degradation. NDN , a p53 target gene, codes for Necdin which was previously shown to regulate the quiescence and response to genotoxic stress of hematopoietic stem/progenitor cells (Asai et al., Blood 2012). Further replication results, an analysis of additive effects and multivariate analyses will be presented.In conclusion, our results demonstrate that germline genetic variation is associated with high levels of MRD after induction/consolidation treatment for ALL and relapse on an AIEOP-BFM ALL protocol. In line with previous studies (Yang et al., JAMA 2009), our data suggest that genome-wide approaches may be useful to identify novel and clinically relevant SNVs for further refinement of risk-adapted treatment strategies in pediatric ALL. DisclosuresSchrappe:JAZZ Pharma: Consultancy, Research Funding; SigmaTau: Consultancy, Research Funding; Medac: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Baxalta: Consultancy, Research Funding.

Bone pain at leukemia diagnosis and other risk factors for symptomatic osteonecrosis in children with acute lymphoblastic leukemia.

Osteonecrosis is a major cause of acute and long-lasting complications of acute lymphoblastic leukemia (ALL) therapy in children. Our study aimed to evaluate the prevalence, characteristics, risk factors, and outcome of osteonecrosis in children with ALL. The cohort included 559 children aged 1-20years diagnosed with ALL between 2003 and 2018 at two tertiary medical centers in Israel and enrolled in two consecutive protocols: ALL-IC BFM 2002 and AIEOP-BFM ALL 2009. Symptomatic osteonecrosis was prospectively captured as an adverse event. Osteonecrosis occurred in 51 patients (9.1%). Ninety-four percent of the events were graded as moderate or severe (grades 3-4, Ponte di Legno Toxicity Working Group classification) and multiple bone involvement was common. Full resolution of osteonecrosis was documented in only 16% of the children (median follow-up 4.2years). Stepwise logistic regression identified five risk factors for osteonecrosis, with a high predictive value (AUC=0.88): older ageat ALL diagnosis, high-risk ALL group, T-cell immunophenotype, female gender, and a novel risk factor: bone pain at the time of leukemia diagnosis. In addition, osteonecrosis was less common among children of Arab ethnicity. Thrombophilia and an elevated age-adjusted body mass index were not confirmed as risk factors for osteonecrosis. Due to the low rates of osteonecrosis resolution and its debilitating long-term impact, the identification of patients at high risk for osteonecrosis is important for their inclusion in further studies evaluating potential therapeutic adjustments.

Hepatic sinusoidal obstruction syndrome and short-term application of 6-thioguanine in pediatric acute lymphoblastic leukemia

Long-term treatment with 6-thioguanine (6-TG) for pediatric acute lymphoblastic leukemia (ALL) is associated with high rates of hepatic sinusoidal obstruction syndrome (SOS). Nevertheless, current treatment continues to use short-term applications of 6-TG with only sparse information on toxicity. 6-TG is metabolized by thiopurine methyltransferase (TPMT) which underlies clinically relevant genetic polymorphism. We analyzed the association between hepatic SOS reported as a serious adverse event (SAE) and short-term 6-TG application in 3983 pediatric ALL patients treated on trial AIEOP-BFM ALL 2000 (derivation cohort) and defined the role of TPMT genotype in this relationship. We identified 17 patients (0.43%) with hepatic SOS, 13 of which with short-term exposure to 6-TG (P < 0.0001). Eight of the 13 patients were heterozygous for low-activity TPMT variants, resulting in a 22.4-fold (95% confidence interval 7.1–70.7; P ≤ 0.0001) increased risk of hepatic SOS for heterozygotes in comparison to TPMT wild-type patients. Results were supported by independent replication analysis. All patients with hepatic SOS after short-term 6-TG recovered and did not demonstrate residual symptoms. Thus, hepatic SOS is associated with short-term exposure to 6-TG during treatment of pediatric ALL and SOS risk is increased for patients with low-activity TPMT genotypes.

Venous Thromboembolism and Its Risk Factors in Children with Acute Lymphoblastic Leukemia in Israel: A Population-Based Study.

Simple SummaryCurrent cure rates of childhood acute lymphoblastic leukemia (ALL) surpass 90% and thus the mitigation of chemotherapy-associated toxicity has become a major goal. Venous thromboembolism (VTE) is a major toxicity of ALL therapy, but its precise prevalence, risk factors, and optimal prophylaxis are yet to be determined. This study evaluated VTE in a cohort of 1191 children, including its rate, risk factors, and long-term sequelae. VTE was found in 7.5% of the cohort, 27% severe events. Most events resolved without late sequelae. We identified four risk factors associated with VTEs: Age >10 years, high-risk ALL group, severe hypertriglyceridemia during therapy, and inherited thrombophilia. Since children with these risk factors had significantly higher rates of severe VTE, a routine evaluation may identify potential candidates for prophylactic interventions.Venous thromboembolism (VTE) is a serious complication of acute lymphoblastic leukemia (ALL) therapy. The aim of this population-based study was to evaluate the rate, risk factors, and long-term sequelae of VTE in children treated for ALL. The cohort included 1191 children aged 1–19 years diagnosed with ALL between 2003–2018, prospectively enrolled in two consecutive protocols: ALL-IC BFM 2002 and AIEOP-BFM ALL 2009. VTEs occurred in 89 patients (7.5%). Long-term sequelae were uncommon. By univariate analysis, we identified four significant risk factors for VTEs: Severe hypertriglyceridemia (p = 0.005), inherited thrombophilia (p < 0.001), age >10 years (p = 0.015), and high-risk ALL group (p = 0.039). In addition, the incidence of VTE was significantly higher in patients enrolled in AIEOP-BFM ALL 2009 than in those enrolled in ALL-IC BFM 2002 (p = 0.001). Severe VTE occurred in 24 children (2%), all of whom had at least one risk factor. Elevated triglyceride levels at diagnosis did not predict hypertriglyceridemia during therapy. In a multivariate analysis of 388 children, severe hypertriglyceridemia and inherited thrombophilia were independent risk factors for VTE. Routine evaluation for these risk factors in children treated for ALL may help identify candidates for intervention.

Acute lymphoblastic leukemia in pediatric epigenetic approach

Introduction and objectives: Acute lymphoblastic leukemia (ALL) in pediatric patients is an issue that affects the quality of life of the patient and his family, so it is urgent to know the physiology, presentation, and functionality of the cell population that allows determining the more effective treatments. The objective is to review the evidence derived from cohort studies and clinical trials on ALL in pediatric patients. Method: A retrospective study carried out based on the search for cohort studies and clinical trials in the last 10 years in MEDLINE, EMBASE, and Cochrane Controlled Trials Register whose keywords [Acute and Lymphoblastic Leukemia], [epigenetical], [Drug Therapy], [Pediatric]. Results: 87 articles were found based on titles and abstracts, of which 16 focus on the age group and criteria of interest. Of the 10% of the known etiology, genetic alterations are more important. However, there are epigenetic modifications that are important for leukemia to occur, such as DNA methylation, histone modification, and regulation by non-coding RNAs. Discussion/conclusions: Acute lymphoblastic leukemia in a heterogeneous pathology and the study of its molecular biology is very useful to identify therapeutic targets and have greater effectiveness in treatment and increase disease-free survival with a longer-term than the current one. In each proposed treatment scheme, it is sought that there be greater effectiveness and less toxicity as in the AIEOP-BFM ALL or IDA-FLAG schemes.