High-risk Acute Lymphoblastic Leukemia Research Articles

Abstract 3271: The casein kinase 2/IKZF1 axis attenuates ceramide accumulation through up-regulation of acid ceramidase: Implications for use of CK2 inhibitors in high-risk pediatric B-cell acute lymphoid leukemia

Abstract Casein Kinase II (CK2), a growth-promoting non-oncogene, is frequently over-expressed in pediatric high-risk B-cell acute lymphoblastic leukemia (B-ALL) leading to proliferation and survival. CK2 phosphorylates the tumor suppressor IKZF1 (IKAROS) resulting in a loss-of DNA-binding activity. Loss of IKZF1 function is associated with poor prognosis in high-risk B-ALL. As such, inhibition of CK2 using silmitasertib (CX-4945), to restore IKZF1 function, is being therapeutically investigated in high-risk B-ALL. CK2 also phosphorylates/activates the mammalian ceramide synthases to increase production of the pro-apoptotic sphingolipid, ceramide (Cer). Therefore, we hypothesized that inhibition of CK2 with CX-4945 would exert both pro-death (restoration of IKAROS function) and pro-survival (decreased Cer production) effects on B-ALL cells. Moreover, this suggests that targeting the sphingolipid metabolic pathway in tandem with CX-4945 could be a novel strategy for treatment of high-risk B-ALL. To this end, we examined the effects of CX-4945 on sphingolipid levels and the enzymes of the sphingolipid metabolic pathway. Unexpectedly, we determined that CX-4945 dose-dependently increased expression of ceramide synthase 1 (CERS1) and acid ceramidase (AC) proteins, in an IKZF1 dependent manner. Increased CERS1 expression/production of C18:0 Cer has been linked to enhanced mitophagy (i.e., lysosomal clearance of damaged mitochondria) in head and neck squamous cell carcinoma. Induction of mitophagy by CX-4945 was confirmed by western blot analysis of mitofusin-1 and Drp-1 levels. Up-regulation of AC, a lysosomal protein, metabolizes C18:0 Cer to sphingosine for recycling and prevents Cer-induced lysosomal membrane permeabilization. Thus, induction of mitophagy and metabolic breakdown of Cer could potentially reduce the apoptotic efficacy of CX-4945 in B-ALL cell lines. We, therefore, tested whether inhibition of AC would synergize with CX-4945 to enhance B-ALL cell death. Indeed, synergy was observed with the AC inhibitor, SACLAC, in multiple cell lines. Consistent with the observation of synergy between CX-4945 and SACLAC, the combination also enhanced cleavage of PARP indicating an enhancement of cell death. We are currently testing the efficacy of a combination of CX-4945 and AC inhibitors in cell-line derived xenograft models of human B-ALL. Together, these findings indicate that simultaneous targeting of CK2 and AC represents a rationally selected combinatorial therapeutic strategy that overcomes the ability of cancer cells to adapt to single agent targeted therapies by altering pro- and/or anti-apoptotic pathways. Citation Format: Jeremy A. Hengst, Diwakar Bastihalli Tukaramrao, Todd Fox, Arati Sharma, Dhimant Desai, Sinisa Dovat. The casein kinase 2/IKZF1 axis attenuates ceramide accumulation through up-regulation of acid ceramidase: Implications for use of CK2 inhibitors in high-risk pediatric B-cell acute lymphoid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3271.

Effect of apolipoprotein E (APOE) gene polymorphisms on the lipid profile of children being treated for acute lymphoblastic leukemia

BackgroundMedications used to treat acute lymphoblastic leukemia (ALL), such as l-asparaginase, can cause blood lipid disturbances. These can also be associated with polymorphisms of the lipoprotein lipase (LpL) and apolipoprotein E (APOE) genes.ProcedureWe aimed to investigate the association between lipid profile, certain LpL and APOE gene polymorphisms (rs268, rs328, rs1801177 and rs7412, rs429358 respectively) as well as the risk subgroup in 30 pediatric patients being treated for ALL, compared with 30 pediatric ALL survivors and 30 healthy controls.ResultsThe only APOE gene polymorphism with significant allelic and genotypic heterogeneity was rs429358. Further analysis of this polymorphism showed that genotype (CC, CT, or TT) was significantly associated with (1) changes in the lipid profile at the end of consolidation (total cholesterol, LDL, apo-B100, and lipoprotein a) and during re-induction (total cholesterol and apo-B100), and (2) classification in the high risk-ALL subgroup (for CC genotype/C allele presence).ConclusionsLipid abnormalities in children being treated for ALL may be associated with the APOE genotype, which is also possibly associated with risk stratification. Further research is needed to confirm the potential prognostic value of these findings.

Abstract P30: Therapeutic activation of pre-B-cell receptor signaling in high-risk pediatric ALL: exploiting TCF3 and MYC-linked vulnerabilities

Abstract P30: Therapeutic activation of pre-B-cell receptor signaling in high-risk pediatric ALL: exploiting TCF3 and MYC-linked vulnerabilities

Peg-asparaginase associated toxicities in children with acute lymphoblastic leukemia: A single-center cross-sectional study

BackgroundLeukemia is the most prevalent type of cancer affecting children, representing approximately 35% of cancer diagnoses in Saudi Arabia. Notably, significant improvement in the overall survival and cure rates for pediatrics with acute lymphoblastic leukemia (ALL) has been attributed to adding asparaginase to the chemotherapy regimen. However, the administration of these agents may lead to a multifactorial range of toxicities that often alter treatment outcomes. ObjectiveThe study aimed to characterize the prevalence of common toxicities related to polyethylene glycol (PEG) asparaginase in children aged 0–14 years diagnosed with ALL. Additionally, it assessed the types of toxicities associated with intravenous (IV) and intramuscular (IM) administration of Peg-asparaginase. MethodIt was an observational retrospective cross-sectional study. Data was extracted from the hospital's electronic health record (EPIC). Using EPIC, we reviewed medical charts of all pediatric patients below 14 years old diagnosed with ALL who received at least one dose of PEG-asparaginase between January 2020 and March 2023. The toxicity grading was based on the National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE). ResultsOne hundred and ninety-one patients with ALL were included in the study. Overall, 79 toxicity episodes were experienced by the study patients. Anaphylaxis/hypersensitivity (36.7%) and hepatotoxicity (31.6%) were the most prevalent toxicities reported, followed by pancreatitis and hyperglycemia (12.7% each). According to the CTCAE grading, approximately 70% of toxicities were categorized as Grade 3 and 4. Notably, 60% of the events occurred during the induction and consolidation phase of therapy. Interestingly, there was no significant difference in rates of toxicities between patients receiving IV or IM PEG-asparaginase. ConclusionThe distribution of toxicities highlighted in our study aligns with findings from previously published studies. Furthermore, multivariate analysis indicated that patients with high-risk ALL and T-ALL were more likely to develop toxicities compared to other forms of ALL.

Biological Markers of High-Risk Childhood Acute Lymphoblastic Leukemia.

Childhood acute lymphoblastic leukemia (ALL) has witnessed substantial improvements in prognosis; however, a subset of patients classified as high-risk continues to face higher rates of relapse and increased mortality. While the National Cancer Institute (NCI) criteria have traditionally guided risk stratification based on initial clinical information, recent advances highlight the pivotal role of biological markers in shaping the prognosis of childhood ALL. This review delves into the emerging understanding of high-risk childhood ALL, focusing on molecular, cytogenetic, and immunophenotypic markers. These markers not only contribute to unraveling the underlying mechanisms of the disease, but also shed light on specific clinical patterns that dictate prognosis. The paradigm shift in treatment strategies, exemplified by the success of tyrosine kinase inhibitors in Philadelphia chromosome-positive leukemia, underscores the importance of recognizing and targeting precise risk factors. Through a comprehensive exploration of high-risk childhood ALL characteristics, this review aims to enhance our comprehension of the disease, offering insights into its molecular landscape and clinical intricacies in the hope of contributing to future targeted and tailored therapies.

Acute Lymphoblastic Leukemia (ALL) as a secondary malignancy in adolescent patient after completion of Ewing's sarcoma chemotherapy: a case report

OBJECTIVE: To report an adolescent patient who was diagnosed with Acute lymphoblastic leukemia (ALL) after completing Ewing sarcoma (ES) therapy. CASE: A 14 years old girl complained of left crusis tumor since 4 months before hospitalized. Tumor biopsy showed a round cell tumor, as well as immunohistochemistry which showed a positive CD99, consistent with ES. The patient had received 14 cycles chemotherapy for 7 months with regimens vincristine 1.5 mg/m2, cyclophosfamide 1200 mg/m2, dactinomycin 0.5 mg/m2, and doxorubicin 30 mg/m2. Between chemotherapy, patient also was given granulocyte colony-stimulating factor several times. Pasien also undergo 31 times radiation therapy with total 31x1.8 Gy (55.8 Gy). Two months after last chemotherapy for Ewing sarcoma, she was admitted again with prolonged fever, and pallor.Laboratorium showed leukocytosis (88.86x103uL), monocytosis (41%), severe neutropenia (0.39x103uL), severe normochromic normocytic anemia (4,3 g/dL), severe thrombocytopenia (4x103uL), reticulocytopenia (0,46%), with lymphoblast in peripheral blood smear. Bone marrow aspiration result showed lymphoblast infiltration 60% with heterogen morphology, no cluster cell non hematopoetic in bone marrow, consisten with acute lymphoblastic leukemia (ALL-L2) suspected secondary malignancy from Ewing’s Sarcoma. Patient underwent chemotherapy for high risk ALL at induction phase with combination of vincristin, daunorubicin, L-asparginase, metotrexate high dose, prednison, and 6-mercaptopurine. After completing induction phase, the patient’s condition get worsening and later died. CONCLUSION: ES survivors are at significant risk for secondary malignancies. Leukimia is one of which the most common secondary malignancies. Young children are particularly susceptible to the mutagenic effects of both chemotherapy and radiotherapy, as proven by several studies.

461 - Management of wound dehiscence during active chemotherapy in a young adult with high risk ALL

461 - Management of wound dehiscence during active chemotherapy in a young adult with high risk ALL

Efficacy and safety in early application of Ruxolitinib for high-risk acute lymphoblastic leukemia to prevent GVHD and recurrence after allogeneic hematopoietic stem cell transplantation

BackgroundAllogeneic hematopoietic stem cell transplantation (allo-HSCT) has been identified as the potentially curative treatment for high-risk acute lymphoblastic leukemia (ALL) in adult patients. However, relapse of the disease and/or development of graft-versus-host disease (GVHD) remain to be the most common barriers for successful allo-HSCT. Preclinical studies showed that ruxolitinib, a Janus tyrosine kinase (Jak)1 and Jak2 inhibitor, has a selective anti-GVHD effects while preserving a potent graft-versus-leukemia (GVL) effect. Our study aimed to investigate the efficacy and safety in early application of ruxolitinib for the high-risk ALL patients to prevent GVHD. MethodsThere were eight patients undergoing allo-HSCT at the Bone Marrow Transplantation Center of the Third Xiangya Hospital of Central South University between April 2020 and April 2021. Ruxolitinib (5–10 mg twice daily) was administered early (median time: 45 days) after stem cell infusion. ResultsAfter a median follow-up of 14 months (range from 8 to 18 months), the ALL disease relapse occurred in two cases. Among all eight patients, two of them developed grade I/II acute (a) GVHD, while no patient developed grade III/IV aGVHD, and one patient developed chronic (c) GVHD. As for the virus activation, no patient developed EBV activation or EBV related lymphoproliferative disease, and three patients developed CMV activation. Our results suggest that the early application of ruxolitinib could safely and effectively prevent the occurrence of GVHD after allo-HSCT for the high-risk ALL patients. However, it may have a limited effect on preventing the recurrence of high-risk ALL and thus may require additional therapy with other anti-relapse drugs. ConclusionsOur preliminary observations suggest that an early application of ruxolitinib can safely and effectively prevent the occurrence of GVHD after allo-HSCT for the high-risk ALL patients. However, ruxolitinib may have a limited effect on preventing the ALL recurrence of high-risk patients.

Fungal Otomastoiditis caused by Aspergillus fumigatus in an immunocompromised patient: case report and literature review

This case report describes a 10-year-old male diagnosed with high-risk B-cell Acute Lymphoblastic Leukemia at a tertiary care center undergoing consolidation with the Total XV protocol. The patient, with a history of Rhino-sinusal Aspergillosis and secondary prophylaxis, presented with recurrent middle ear infections, bilateral non-purulent serous discharge, otalgia, mastoid pain, persistent high-grade fever, and profound neutropenia. Fungal ostomastoiditis was diagnosed and treated with targeted antifungals, yielding a positive response.

Invasive Fungal Infections in Children with Acute Leukemia: Epidemiology, Risk Factors, and Outcome.

Invasive fungal infections (IFI) cause morbidity and mortality in children with acute leukemia (AL). We retrospectively collected data on febrile neutropenic episodes (FNE) in AL children (2016-2021) and assessed factors associated with proven/probable IFI. Ninety-three children developed 339 FNE. Seventeen (18.3%) children developed 19 proven/probable IFI (11 yeast; eight molds). The proven/probable yeast IFI rate was 6/52 (11.5%) in children who belong to the high risk for IFI category (HR-IFI-AL: high-risk acute lymphocytic leukemia (ALL), acute myeloid leukemia, relapse); and 5/41 (12.2%) in the non-HR-IFI-AL category (standard/intermediate risk ALL). The proven/probable mold IFI rate was 7/52 (13.5%) in HR-IFI-AL children and 1/41 (2.4%) in the non-HR-IFI-AL category. In the multivariable analysis, underlying genetic syndrome, oral mucositis, and older age were significantly associated with proven/probable IFI, while a longer time since AL diagnosis was protective. Two of 13 (15.4%) HR-IFI-AL children died because of IFI. The elevated risks of proven/probable mold IFI and the associated mortality in HR-IFI-AL children, and high risk of invasive candidiasis in the non-HR-IFI-AL group, emphasize the need for the close monitoring of local epidemiology and the adjustment of practices accordingly.

Comparison of high-throughput single-cell RNA-seq methods for ex vivo drug screening.

Functional precision medicine (FPM) aims to optimize patient-specific drug selection based on the unique characteristics of their cancer cells. Recent advancements in high throughput ex vivo drug profiling have accelerated interest in FPM. Here, we present a proof-of-concept study for an integrated experimental system that incorporates ex vivo treatment response with a single-cell gene expression output enabling barcoding of several drug conditions in one single-cell sequencing experiment. We demonstrate this through a proof-of-concept investigation focusing on the glucocorticoid-resistant acute lymphoblastic leukemia (ALL) E/R+Reh cell line. Three different single-cell transcriptome sequencing (scRNA-seq) approaches were evaluated, each exhibiting high cell recovery and accurate tagging of distinct drug conditions. Notably, our comprehensive analysis revealed variations in library complexity, sensitivity (gene detection), and differential gene expression detection across the methods. Despite these differences, we identified a substantial transcriptional response to fludarabine, a highly relevant drug for treating high-risk ALL, which was consistently recapitulated by all three methods. These findings highlight the potential of our integrated approach for studying drug responses at the single-cell level and emphasize the importance of method selection in scRNA-seq studies. Finally, our data encompassing 27327 cells are freely available to extend to future scRNA-seq methodological comparisons.

Risk factors and clinical impact of thrombosis during induction chemotherapy for pediatric acute lymphoblastic leukemia: A report from CYP-C.

Thromboembolism (TE) is associated with reduced survival in pediatric acute lymphoblastic leukemia (ALL). It has been hypothesized that TE might signal leukemic aggressiveness. The objective was to determine risk factors for TE during ALL induction (TEind ) therapy and whether TEind is associated with treatment refractoriness. This retrospective cohort study using the population-based Cancer in Young People Canada (CYP-C) registry included children <15 years of age diagnosed with ALL (2000-2019) and treated at one of 12 Canadian pediatric centers outside of Ontario. Univariate and multivariable logistic regression models were used to determine risk factors for TEind and whether TEind predicted induction failure and ALL treatment intensification. The impact of TEind on overall and event-free survival was estimated using Cox proportional hazard regression models. The study included 2589 children, of which 45 (1.7%) developed a TEind . Age (<1 year and ≥10 years vs. 1-<10 years), T-cell phenotype, high-risk ALL, and central nervous system involvement were all associated with TEind in univariate analysis. Age and T-cell phenotype remained independent predictors of TEind in multivariable analysis. Induction failure occurred in 53 patients (2.1%). TEind was not associated with induction failure (OR: not estimable) or treatment intensification (adjusted OR [95% CI]: 0.66 [0.26-1.69]). TEind was independently associated with overall survival (adjusted HR [95% CI]: 2.54 [1.20-5.03]) but not event-free survival (adjusted HR [95% CI] 1.86 [0.98-3.51]). In this population-based study of children treated with contemporary chemotherapy protocols, TEind was associated with age and T-cell phenotype and mortality but did not predict induction failure.

Intensified conditioning regimens with total marrow irradiation/etoposide/cyclophosphamide and busulfan/etoposide/cyclophosphamide overcome the impact of pre-transplant minimal residual disease on outcomes in high-risk acute lymphoblastic leukemia patients in complete remission.

Among high-risk acute lymphoblastic leukemia (ALL) patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT), those with positive minimal residual disease (MRD) are susceptible to poor outcomes. Therefore, it is necessary to determine the most suitable preparatory regimen for these patients. Data were analyzed from 141 patients who received allo-HSCT and were diagnosed with high-risk ALL. These patients underwent intensified conditioning regimens, including either total marrow and lymphoid irradiation (TMLI)-etoposide (VP16)-cyclophosphamide (CY) or busulfan (BU)-VP16-CY between October 2016 and November 2022. A total of 141 individuals were in complete remission (CR) before transplantation and, among all patients, 90 individuals exhibited a negative MRD status and 51 patients had a positive MRD status. In patients who tested negative for MRD, the incidence of relapse within a 2-year timeframe was 25.0% (24.8%-25.5%), compared with 32.2% (31.2%-33.2%) in MRD-positive patients; however, this difference was not statistically significant. There were no significant differences in the 2-year disease-free survival (DFS) and 2-year overall survival (OS) rates between the MRD-negative and MRD-positive groups (DFS: 67.2% (57.9%-78.1%) vs. 55.5% (42.6%-72.3%); OS: 69.0% (61.9%-88.2%) vs. 66.7% (53.9%-82.5%)). Furthermore, no notable variations were observed in the occurrence of transplant-related mortality (TRM) and graft-versus-host disease (GVHD) across the two groups. This study reveals the benefits of TMLI-VP16-CY and BU-VP16-CY conditioning regimens in high-risk ALL patients with CR and MRD-positive status. A large-scale prospective clinical trial is warranted in the future.

ALCL Protocol of the DOIG Research Institute 2003: results of treatment of anaplastic large cell lymphoma in children in a comparative multicenter clinical trial

Introduction. Anaplastic large cell lymphoma (ALCL) is characterized by clinical, morphological and immunohistochemical heterogeneity. Both intensive block regimens and programs similar to those used in the treatment of acute lymphoblastic leukemia (ALL) are used in therapy. Taking into account the prognostically unfavorable effect of the expression of T-cell markers by tumor cells, a protocol ALCL NII DOIG 2003 was developed, which takes into account not only risk groups, but also immunophenotypic features of the tumor substrate.Aim. To evaluate the treatment effectiveness of children with ALCL according to the protocol ALCL NII DOIG 2003 in comparison with the standard protocol NHL-BFM 95.Materials and methods. The study included 100 patients with newly diagnosed ALCL who received treatment from 2000 to 2023 in 5 federal and regional pediatric oncohematology departments. The patients were divided into 2 groups depending on the protocol of therapy: I group — 48 patients treated with protocol ALCL NII DOIG 2003; II group — 52 patients treated according to the NHL-BFM 95 protocol. The basis of the protocol ALCL NII DOIG 2003 was made up of high-intensity block regimes similar to those used in the treatment of high-risk T-precursor ALL. A comparative assessment of the overall, event-free and relapse-free survival of patients was carried out depending on the therapeutic protocol using the SPSS 21.0 program. Results. The 10-year overall survival rate in patients from I group was 95.3 ± 3.3 %, II group — 82.0 ± 5.4 % (p = 0.037). 10- year event-free survival in I group was 95.3 ± 3.3 %, II — 68.6 ± 6.5 % (p = 0.001). 10-year relapse-free survival in I group of patients was 97.3 ± 2.7 %, while in the II group — 74.4 ± 6.4 % (p = 0.003).Conclusion. The obtained results indicate the high effi ciency of a differentiated, immuno-oriented approach to the treatment of ALCL with the ALCL NII DOIG 2003 protocol, making it possible to achieve signifi cantly higher survival rates of patients compared to the standard treatment protocol.

Outcomes of allogeneic stem cell transplantation in acute leukemias – single centre study

Aim: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is recommended in first complete remission (CR) for adverse and intermediate-risk acute myeloid leukemia (AML) and not recommended for AML patients with favourable disease. Allo-HSCT is standard of care in high-risk acute lymphoblastic leukaemia (ALL) and relapsed ALL but is not indicated for standard-risk ALL. Even though in many studies benefit for survival was proven for these patients, relapse rates (RR) at one year (32.9% for LAM and 34% for LAL) remains problematic. We want to report the outcomes of patients treated for acute leukaemia in our centre. Materials and methods: We conducted a retrospective descriptive analysis of 215 patients with acute leukaemia treated with allo-HSCT between 2017 and 2022 in Fundeni Clinical Institute. Data analysis aimed the overall survival (OS) rate, progression free survival (PFS) and RR at one year based on genetic risk, number of CR, minimal residual disease (MRD) status and conditioning regimen used. Results: At one year evaluation we observed OS rate of 64% and a 34% RR for ALL patients, while AML patients had a 74% OS rate and 13% RR. Conclusions: The results obtained in our centre are similar to the ones reported in literature. The highest risk of relapse and smallest rate of survival are registered for patients who present with negative risk factors. The multifactorial influence over OS and RR impose the need for future studies in which different patient categories can benefit from an optimal therapeutic management.

Abstract C055: Distinct epigenomic regulation of gene expression in Hispanic/Latino children with high-risk B-cell acute lymphoblastic leukemia

Abstract B-Cell Acute Lymphoblastic Leukemia (B-ALL) is the most common childhood malignancy. Hispanic/Latino (H/L) children have a 40% higher death rate than non-Hispanic white (NHW) children after correcting for socioeconomic factors. Recently, we identified a 3.5-fold increased incidence of deletion of one IKZF1 allele (IKZF1-del) and over 15-fold increased incidence of deletion of one IKZF1 allele concomitant with IGH-CRLF2 translocation (IKZF1-del/IGH-CRLF2) in B-ALL of H/L children. This provided a biological rationale for the worse prognosis of B-ALL in H/L children. The very high incidence (29% in all H/L children and 60% in ≥10yr old H/L children) makes IKZF1-del the most frequent genetic alteration that confers adverse prognosis in B-ALL in H/L children. The IKZF1 gene encodes a DNA-binding protein, IKAROS, which regulates transcription of its target genes via chromatin remodeling. The purpose of the study is to analyze the global epigenomic regulation of gene expression in H/L and NHW children with B-ALL and identify biological determinants of altered gene expression that result in worse prognosis for H/L children with B-ALL. /. B-ALL primary cells from 60 H/L and NHW children were analyzed with whole genome sequencing and gene expression profiling using RNA-seq. A subset of B-ALL from each group was analyzed for epigenomic changes by Reduced Representation Bisulfite Sequencing (RRBS), ChIP-seq, CUT&amp;RUN, and CUT&amp;Tag. Results showed that gene expression pattern in B-ALL of H/L children is quite different from B-ALL of NHW children. GSEA and Ingenuity pathway analysis identified oncogenic drivers of B-ALL in H/L children. B-ALL of H/L children have elevated expression of genes that promotestemness, inflammation, and with antiapoptotic activity. RBBS analysis showed a distinct DNA methylation pattern in B-ALL in H/L vs NHW children at promoters of a large set of genes that are involved in cellular signaling, RNA processing, and cellular proliferation. Presence of IKZF1-del results in the alteration of global epigenomic regulation of gene expression with altered enhancer and super-enhancer landscape. Epigenomic alterations due to IKZF1-del resulted in profound redistribution of enhancers and super-enhancers in B-ALL of both H/L vs. NHW children. This was associated with a distinct alteration in gene expression and uncovered that IKAROS regulates different sets of genes in B-ALL in H/L vs. NHW children. These data identified a unique oncogenic signaling that are responsible for aggressive course of B-ALL in H/L children. In conclusion, the multiomics analysis of primary B-ALL cells revealed a distinct global epigenomic regulation of gene expression between B-ALL from H/L and NHW children. This was demonstrated by alterations in DNA methylation and enhancer landscape. Presence of IKZF1-del has a different effect on gene expression in B-ALL of H//L vs NHW children. Results of the study provide a biological rationale for the worse prognosis of B-ALL in H/L children and identified novel therapeutic targets for this disease. Citation Format: Laura Budurlean, Bing He, Yali Ding, Diwakar Bastihalli Tukaramrao, Chingakham Singh, Joseph Schramm, Daniel Bogush, James Broach, Sinisa Dovat. Distinct epigenomic regulation of gene expression in Hispanic/Latino children with high-risk B-cell acute lymphoblastic leukemia [abstract]. In: Proceedings of the 16th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2023 Sep 29-Oct 2;Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2023;32(12 Suppl):Abstract nr C055.

Phenotypic Drug Response Profiling Identifies Asparaginase-Based Synergistic Combinations for Very High Risk Acute Lymphoblastic Leukaemia

In acute lymphoblastic leukaemia (ALL), empirical multi-drug chemotherapy combinations lead to &amp;gt;85% cure rates in children. But 10-15% of patients show suboptimal response and as drugs are dosed to tolerance, further intensification is not possible. An exception is the key antileukaemic drug, asparaginase (ASNase), which is not associated with dose-related toxicity. We evaluated the use of ex-vivo co-culture based drug response profiling (DRP) to identify novel synergistic and effective drug combinations with ASNase for patients with suboptimal responses to standard chemotherapy in ALL. Patient-derived xenograft (PDX) ALL cells from 5 patients with high-risk B-ALL (3 B-other, 2 IKZF plus) were treated with 120 drugs in 5-point serial dilutions in combination with or without an IC 50 dose of ASNase, in co-cultures with hTERT-immortalized primary bone marrow stromal cells. Eight clinically relevant agents with promising sensitizing activity as identified by decreased area-under-the-curve with addition of ASNase were identified (SEL, selinexor; ELTA, eltanexor; VEN, venetoclax; BZM, bortezomib; navitoclax; carfilzomib; mitoxantrone, birinapant). We validated combinations of ASNase with the 8 selected drugs plus 4 drugs used in induction therapy (prednisolone, dexamethasone, vincristine and daunorubicin) using a 4x4 drug matrix in 15 high-risk ALL PDX samples. Synergy scores were calculated using the zero-interaction potency (ZIP) model. Synergy (ZIP score ≥10) or additivity (ZIP score &amp;gt;0 and &amp;lt;10) with ASNase was identified for exportin-1 inhibitors (XPO1i; selinexor, eltanexor), venetoclax and proteosomal inhibitors (bortezomib, carfilzomib) in 100% (15/15), 86.6% (13/15) and 80-86.6% (12/15 and 13/15) samples respectively. Antagonism (ZIP score &amp;lt;0) was noted in more than one-third (6/15) of samples with ASNase in combination with steroids or vincristine. Three-drug combinations were evaluated ex-vivo in these 15 PDX samples. Synergy was identified with addition of low doses of BZM or VEN to the ASNase-SEL combination (ASNase-SEL-BZM and ASNase-SEL-VEN). Next, 4-6 mice per treatment arm were transplanted intravenously with 1 million ALL cells from 4 high-risk B-ALL patients. Randomized cohorts were treated with vehicle, ASNase-BZM-ELTA (ABE) and ASNase-dexamethasone-vincristine (ADV). Leukemic burden was assessed weekly using flowcytometry. The mice tolerated both treatment arms well. Platelet counts recovered within one week after an initial drop in the ABE arm. Both treatment groups had significant improvement in event free survival (EFS) as compared to the control group (ABE p-value 0.0082; ADV p-value 0.0325) though no significant differences were noted between the 2 treatment arms (p-value 0.8971). Three patients with high-risk medullary relapsed/refractory BCP-ALL, 2 not in morphological remission and 1 at time of very early relapse, received a combination of ASNase-BZM-VEN-SEL (ABVS) after consent. One patient for whom DRP suggested sensitivity to both SEL and VEN achieved molecular remission (molecular minimal residual disease &amp;lt; 10 -5) after one course of therapy remaining in complete remission 11 months later. Two patients with sensitivity to SEL but not to VEN as assessed by DRP, had progressive disease. ABVS was well tolerated in these patients with no grade 3-4 adverse events. Our results identify XPO1 inhibitors as a new class of drugs with promising anti-leukaemic activity in BCP-ALL. Co-culture based DRP shows promise in identifying alternative novel sensitive synergistic combinations ex-vivo with decreased toxicity for patients who have an inadequate response to standard therapy.

Experience with Blinatumomab in Pediatric Patients in Argentina. Multi-Center Study

Introduction. Very high-risk and relapsed pediatric acute lymphoblastic leukemia (ALL) require high-dose chemotherapy and hematopoietic stem cell transplant (HSCT) as consolidation of treatment. Patients with positive minimal residual disease (MRD) previous to HSCT have lower survival rates. Immunotherapy is thus useful for management of MRD previous to HSCT with less chemotherapy-associated morbidity. Blinatumomab (Blina) is a bi specific monoclonal antibody against CD19 and CD3ε. Objectives. To report the experience with Blina in children with B-ALL in Argentina and assess its toxicity, effectiveness and overall survival (OS). Patients and methods. Retrospective analytical study. The main Argentinian pediatric hospitals participated. Patients younger than 17 years old with B-ALL in 1 st, 2 nd or 3 rd CR who had received 1 or 2 cycles of Blina were included. Those with negative MRD and those who didn´t complete a cycle were excluded. Toxicity was evaluated according to CTC 4.0 and effectiveness according to pre and post-infusion MRD. OS was calculated using Kaplan Meier with GraphPad Prism V5.0. Results. Patients. Thirty patients received Blina and 6 were excluded: 3 due to negative MRD, 2 didn´t complete 1 cycle and 1 for both reasons.Eligible: 24 patients. The median age was 5.5 (range 0.8-17) years. Patients under 1 year: 3. Distribution by sex F/M= 1/1.8. Patients with High Risk ALL represented 42% (n=10) and relapsed ALL 58% (n=14), 10 in 1 st relapse and 4 in 2 nd relapse. Sixteen patients received 1 cycle and 8 received 2. Median MRD previous to Blina was 0.71 (range 0.023-49)%. Thirteen patients received it in CR (MRD &amp;lt;1%): 7 in 1 st CR, 4 in 2 nd CR, and 2 in 3 rd CR. MRD was 1-5% in 6 patients and &amp;gt;5% in 5. Toxicity. During the 1 st and 2 nd weeks, 25% (n=6) and 16% (n=4) presented GII-III neurological toxicity respectively. Blina infusion was temporarily withdrawn for 3 hours to 12 days in 6 patients but all completed the cycle. During the 1 st week, 41.5% (n=10) presented fever. During the entire 1 st cycle 25% (n=6) presented GIII-IV hematological toxicity, 2 patients presented GIV and 1 GIII infectious toxicity. Regarding the 8 patients who received a 2 nd cycle, 1 patient presented febrile neutropenia and 2 presented GIII infectious toxicity without evidence of neurological toxicity. Systemic inflammatory response syndrome was not evidenced. There were no fatal events related to Blina administration. Efficacy. After the 1 st cycle of Blina 58% became MRD negative (n=14), 17% remained in CR (n=4) and 25% didn´t show any response. All patients who received a 2 nd cycle were in CR; however, 5 of them became MRD negative and 3 relapsed later. Ten of the 13 patients who received Blina in CR with MRD+ became negative and 3 relapsed. Of the 5 with MRD 1-5%, 4 became negative, 1 achieved CR with MRD+ and 1 progressed. Of the 5 patients with MRD&amp;gt;5%, 2 achieved CR but none became MRD negative. Fifty percent of the patients received HSCT (n=12): 6 haploidentical, 2 related allogeneic and 4 unrelated. With a median follow-up of 3.5 (1.7-56.9) months, OS at 36 months was 20.3%. Nine patients are alive and in CR (37.5%), 6 post-HSCT and 3 post-Blina still receiving treatment. Five died in CR (4 due to HSCT complications) and 10 died due to disease relapse/progression (3 post HSCT and 7 post-Blina). Conclusions. Blina infusion was safe with the previously described toxicities. The highest response rate was evidenced in patients in CR or with MRD 1-5% pre-Blina. The low survival rate in this cohort may be due to the high rate of relapse/progression and toxicity associated with HSCT, although HSCT is essential to consolidate Blina´s response. Blina therapy is effective in patients with very high risk or relapsed ALL with +MRD. The strategy should focus on the precise indication of patients with MRD+ as a bridge to HSCT.

Venetoclax+Azacytidine Followed By Modified BuCy Conditioning Regimen for High Risk or Refractory/Relapsed Acute Lymphoblastic Leukemia and High Risk Myelodysplastic Syndromes

Background:Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only potential curable treatment for myelodysplastic syndromes (MDS) and acute lymphoblastic leukemia (ALL). However, for patients with high-risk or refractory relapsed (R/R) B-ALL with allo-HSCT, the three-year overall survival (OS) rate is only 5-10%, more worse in T-ALL. It is widely known that MDS patients in the high-risk group do not benefit from induction chemotherapy, the five-year OS rate is 23-39% when they received allo-HSCT. Nevertheless, relapse remains a significant cause of treatment failure after allo-HSCT, highlighting the urgent need for improved conditioning regimens that can improve prognosis and reduce relapse rates. Azacytidine (AZA) combined with Venetoclax (VEN), showed a synergistic anti-tumor activity against several hematological malignancies, Therefore, we explored AZA+VEN (VA) combined with modified BuCy(mBuCy) (semustine, cytarabine, busulfan, and cyclophosphamide) conditioning regimen and assessed the safety and effectiveness. Aims ：The aim of this study is to assess the safety, and effectiveness of the VA treatment regimen followed by mBuCy as conditioning regimen for high-risk or R/R ALL(NCT05809167) and high-risk MDS(NCT03256071) . Methods:Patients diagnosed with high-risk or R/R ALL or high-risk MDS (IPSS-R score), undergoing VA+mBuCy as conditioning regimen, and consolidation treatment with demethylating drugs every three months after transplantation, for a total of 8-12 cycles. Results:Patients diagnosed with high-risk or R/R ALL(n=11) or high-risk MDS (IPSS-R score)(n=8) were enrolled between January 13, 2022, and June 9, 2023. The ALL cohort comprised of 5 Ph-negative B-ALL, 2 Ph-positive B-ALL, 3 mixed phenotype leukemias (MPAL), and one T-lymphoblastic lymphoma/leukemia (T-ALL/LBL). There were 7 males and 4 females, with a median age of 33 years (17-54 years) (Table 1). Out of the eleven patients, four had previously undergone Chimeric Antigen Receptor T-Cell Immunotherapy (CAR-T) therapy, and one patient had received Blinatumomab. One patient experienced relapse, another patient was refractory, and 3 patients had extramedullary infiltration. All patients achieved morphological complete remission prior to transplantation, and 10 patients (90.9%) achieved a MRD negative remission. Additionally, all the transplantations successfully completed, with a median time to absolute neutrophil counts (ANC) recovery of 12 days (11-15 days) and platelet (PLT) recovery of 16 days (15-21 days). The median follow-up time was 6.4 months (2.4 -13.6 months), no patient experienced relapse or death, the OS and DFS are both 100%. There were no grade 3 or higher adverse events (AEs), hepatic veno-occlusive disease (VOD),and the transplant-related mortality (TRM). Four patients who developed acute graft-versus-host disease (aGVHD) post-transplantation, all cases involved grade I/II skin aGVHD, and one of them involved grade III GI aGVHD, and no chronic graft-versus-host disease (cGVHD) were observed during the follow-up period. By the endpoint of follow-up, Cytomegalovirus (CMV) activation was 45.4% (5/11) and Epstein-Barr virus (EBV) activation was 27.3% (3/11). The MDS cohort consisted of 7 cases of MDS-EB-I, 1 case of MDS-EB-II. There were 5 males and 3 females, with a median age of 42 years (23-57 years)(Table 2). Two patients received chemotherapy before transplantation, and the other six patients directly received allo-HSCT. In MDS cohort, the median recovery time for ANC was 12 days (11-13 days) and PLT was 17.5 days (16-20 days). The median follow-up was 14.3 months (1.2-18.3 months), no patient had relapsed or died, both OS and DFS are 100%. There were no grade 3 or higher AEs, and TRM. One patient developed VOD. Three patients developed aGVHD after transplantation, all involving degree I/II skin aGVHD, and there was one case of cGVHD during the follow-up period, where the patient developed a mild pulmonary cGVHD. By the endpoint of the follow-up, the rate of CMV activation was 37.5% (3/8) and EBV activation was 25% (2/8). Conclusion:The VA combined with the mBuCy proves to be a effective and safe conditioning regimen for high-risk ALL and MDS patients. However, more patients are needed for follow-up and validation.

Allo-HSCT Improved Outcome of High Risk ALL in Pediatric-Inspired PDT-ALL-2016 Cohort

Background Pediatric-inspired regimen have remarkably improved the prognosis of adult ALL, particularly for adolescent and young adult (AYA) patients. The role of allo-HSCT was controversy in the era of pediatric-inspired regimen. Some study revealed that patients treated with pediatric-inspired regimen those who achieved MRD negative may not require allo-HSCT (Ref1. Ribera et.al, Blood 2021; Ref2. Dhedin et.al, Blood, 2015). To address this issue, we performed study on HR ALL population in pediatric-inspired PDT-ALL-2016 cohort, by comparing outcome of allo-HSCT and chemotherapy groups. Methods PDT-ALL-2016 cohort is a GRAALL-backbone pediatric-inspired regimen (NCT03564470), patients diagnosed with ALL in Nanfang hospital were enrolled in JAN-2016 to DEC-2021. High-risk features included: WBC higher than 30×10^9/L for B-ALL or 100×10^9/L for T-ALL; pro-B or ETP; presence of t(9;22), t(1;19), t(4;11) or 11q23 rearrangements, Ph-like, complex karyotype, hypodiploid or near triploid; IKZF1 deletion/muation. Patients diagnosed with HR-ALL and achieved CR1 after induction were enrolled in this study. The left-truncated Kaplan-Meier estimator was used to compare the overall survival (OS), event-free survival (EFS), cumulative incidence of relapse (CIR) and non-relapse mortality (NRM). Results A total of 265 HR-ALL patients were enrolled in this study, 168 patients in allo-HSCT and 97 patients in chemotherapy cohorts, respectively. The baseline characteristics between two cohorts were comparable. With a median follow-up time of 43.6 months (3.8-60.0 months), the 3-yr OS and EFS were better in allo-HSCT cohort compared with chemo-only cohort (3-yr OS, 76.8% VS 55.6%, P＜0.001; 3-yr EFS, 66.5% VS 38.8%, P＜0.001). Meanwhile, the allo-HSCT cohort had lower CIR and higher but acceptable NRM compared with chemo-only cohort (3-yr CIR, 15.5% VS 55.5%, P＜0.001; 3-yr NRM, 13.8% VS 3.2%, P＜0.001; Table 1). For MRD positive subgroup (allo-HSCT, n=64; chemo-only, n=37), patients who underwent allo-HSCT had longer EFS and OS, and lower CIR compared with chemo-only cohort (3-yr OS, 71.0% VS 35.3%, P＜0.001; 3-yr EFS, 59.3% VS 25.6%, P＜0.001; 3-yr CIR, 19.8% VS 69.8%, P＜0.001). Of note, for MRD negative subgroup (allo-HSCT, n=96; chemo-only, n=51), the allo-HSCT cohort had longer EFS and lower CIR compared with chemo-only cohort, the OS also improved by allo-HSCT (3-yr OS, 81.2% VS 71.0%, P=0.173; 3-yr EFS, 71.1% VS 50.0%, P=0.014; 3-yr CIR, 14.6% VS 48.8%, P＜0.001; Fig 1). In multivariable analyses, allo-HSCT was protective factors for OS (HR=0.37), EFS (HR=0.56) and CIR (HR=0.45), and negatively infected NRM (HR=4.66). Conclusion Here, we showed that allo-HSCT further improved the outcome of HR ALL regardless of the MRD status, in the era of pediatric-inspired regimen. The survival advantage of allo-HSCT came from lower CIR compared with chemotherapy only. Furthermore, compared with published data, the NRM was more acceptable in our cohort. It is the high NRM (20-40%) in other studies ( Ref1. Ribera et.al, Blood 2021; Ref2. Dhedin et.al, Blood, 2015; Ref3. Wieduwilt et.al, leukemia, 2021; Ref4. Seftel et.al, AJH, 2015) that counterbalanced the benefit of CIR by allo-HSCT, which highlight the importance of experienced management for total therapy in adult ALL. This research was supported by the National Natural Science Foundation of China(NFSC82170163, 81970147), Clinical Study of Nanfang Hospital(LC2016ZD009/2019CR012).