Relapsed Childhood Acute Lymphoblastic Leukemia Research Articles

The outcome of relapsed childhood acute lymphoblastic leukemia after allogeneic hematopoietic stem-cell transplantations: A single-center experience.

In children with high-risk childhood acute leukemia who undergo allogeneic hematopoietic stem-cell transplantation (allo-HSCT), relapse is still the leading cause of treatment failure. The prognosis is poor, yet prospective studies have only limited data on risk factors and outcomes. We aimed to understand the outcomes and prognostic factors for patients with acute lymphoblastic leukemia (ALL) who relapsed following allo-HSCT. We analyzed retrospectively 46 children with childhood acute lymphoblastic leukemia who had relapsed after receiving their first alloHSCT. All these patients received salvage chemotherapy which consisted of fludarabine, cytarabine, and idarubicin before performing a second alloHSCT. The median follow-up of the 46 patients after the first transplantation was 366 days. The median time from first allo-HSCT to relapse was 278.4±238.4 days. Forty-six patients received salvage chemotherapy before the second alloHSCT, and CR was achieved in 32 of 46 patients. However, only 17 (37%) of 46 patients received a second allo-HSCT, and 15 of 46 patients died from disease progression, infections, and bleeding. Twelve patients are still alive after the second allo-HSCT. Two-year overall survival (OS) was 38.9%. Local therapy was given to 10 (21.8%) patients, either as part of systemic therapy or alone. In multivariate analyses, the time of relapse and curative salvage therapy with a second allo-HSCT were identified as significant prognostic factors for OS. Children with leukemia who had relapsed after the first allo-HSCT received salvage chemotherapy. Our statistical analysis showed that the second HSCT could be beneficial for outcomes if patients relapsed beyond 180 days of the first allo-HSCT.

Application of prime editing system to introduce TP53 R248Q hotspot mutation in acute lymphoblastic leukemia cell line.

In childhood acute lymphoblastic leukemia (ALL), TP53 gene mutation is associated with chemoresistance in a certain population of relapsed cases. To directly verify the association of TP53 gene mutation with chemoresistance of relapsed childhood ALL cases and improve their prognosis, the development of appropriate human leukemia models having TP53 mutation in the intrinsic gene is required. Here, we sought to introduce R248Q hotspot mutation into the intrinsic TP53 gene in an ALL cell line, 697, by applying a prime editing (PE) system, which is a versatile genome editing technology. The PE2 system uses an artificial fusion of nickase Cas9 and reverse-transcriptase to directly place new genetic information into a target site through a reverse transcriptase template in the prime editing guide RNA (pegRNA). Moreover, in the advanced PE3b system, single guide RNA (sgRNA) matching the edited sequence is also introduced to improve editing efficiency. The initially obtained MDM2 inhibitor-resistant PE3b-transfected subline revealed disrupted p53 transactivation activity, reduced p53 target gene expression, and acquired resistance to chemotherapeutic agents and irradiation. Although the majority of the subline acquired the designed R248Q and adjacent silent mutations, the insertion of the palindromic sequence in the scaffold hairpin structure of pegRNA and the overlap of the original genomic DNA sequence were frequently observed. Targeted next-generation sequencing reconfirmed frequent edit errors in both PE2 and PE3b-transfected 697 cells, and it revealed frequent successful edits in HEK293T cells. These observations suggest a requirement for further modification of the PE2 and PE3b systems for accurate editing in leukemic cells.

TBI, etoposide, and cyclophosphamide conditioning for intermediate-risk relapsed childhood acute lymphoblastic leukemia.

In children with intermediate-risk relapsed acute lymphoblastic leukemia (ALL), allogeneic hematopoietic stem cell transplantation (allo-HSCT) has markedly improved the outcome of patients with an unsatisfactory minimal residual disease (MRD) response. Total body irradiation (TBI), etoposide (ETP), and cyclophosphamide (CY) have been shown to be equivalent to or better than TBI + ETP for conditioning, so we hypothesized that even greater survival could be achieved due to recent advances in HSCT and supportive care. We prospectively analyzed the efficacy and safety of allo-HSCT with a unified conditioning regimen of TBI + ETP + CY in children with intermediate-risk relapsed ALL, based on MRD in the bone marrow after induction, from the Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG) ALL-R08-II nationwide cohort (UMIN000002025). Twenty patients with post-induction MRD ≥ 10-3 and two not evaluated for MRD underwent allo-HSCT. Engraftment was confirmed in all patients, and no transplantation-related mortality was observed. The 3-year event-free survival and overall survival rates after transplantation were 86.4% ± 7.3% and 95.5% ± 4.4%, respectively. Allo-HSCT based on post-induction MRD with TBI + ETP + CY conditioning was feasible in Japanese children with intermediate-risk relapsed ALL.

PPARG, GNG12, and CD19 are potential independent predictors of central nerve recurrence in childhood acute lymphoblastic leukemia

ABSTRACTObjectiveTo identify biomarkers that can predict the recurrence of the central nervous system (CNS) in children with acute lymphoblastic leukemia (ALL).Materials and MethodsThe transcriptome and clinical data of ALL in children were downloaded from the TARGET database. Transcriptome data were analyzed by bioinformatics method to identify core (hub) genes and establish a risk assessment model. Univariate Cox analysis was performed on each clinical data, and multivariate Cox regression analysis was performed on the obtained results and risk score. The children ALL phase I samples from TARGET database were used for validation.ResultsUnivariate multivariate Cox analysis of 10 hub genes identified showed that PPARG (HR = 0.78, 95%CI = 0.67–0.91, p = 0.007), CD19 (HR = 1.15, 95%CI = 1.05–1.26, p = 0.003) and GNG12 (HR = 1.25, 95%CI = 1.04–1.51, p = 0.017) had statistical differences. The risk score was statistically significant in univariate (HR = 3.06, 95%CI = 1.30–7.19, p = 0.011) and multivariate (HR = 1.81, 95%CI = 1.16–2.32, p = 0.046) Cox regression analysis. The survival analysis results of the high and low-risk groups were different when the validation dataset was substituted into the model (p = 0.018). Then, we constructed a Nomogram which had a concordance index of survival prediction of 0.791(95%CI= 0.779-0.803). In addition, the CNS involvement grading status at first diagnosis CNS3 vs. CNS1 (HR = 5.74, 95%CI = 2.01–16.4, p = 0.001), T cell vs B cell (HR = 1.63, 95% CI = 1.06-2.49, p = 0.026) were also statistically significant.ConclusionsPPARG, GNG12, and CD19 may be predictors of CNS relapse in childhood ALL.

Targeting Glutaminolysis Shows Efficacy in Both Prednisolone-Sensitive and in Metabolically Rewired Prednisolone-Resistant B-Cell Childhood Acute Lymphoblastic Leukaemia Cells

The prognosis for patients with relapsed childhood acute lymphoblastic leukaemia (cALL) remains poor. The main reason for treatment failure is drug resistance, most commonly to glucocorticoids (GCs). The molecular differences between prednisolone-sensitive and -resistant lymphoblasts are not well-studied, thereby precluding the development of novel and targeted therapies. Therefore, the aim of this work was to elucidate at least some aspects of the molecular differences between matched pairs of GC-sensitive and -resistant cell lines. To address this, we carried out an integrated transcriptomic and metabolomic analysis, which revealed that lack of response to prednisolone may be underpinned by alterations in oxidative phosphorylation, glycolysis, amino acid, pyruvate and nucleotide biosynthesis, as well as activation of mTORC1 and MYC signalling, which are also known to control cell metabolism. In an attempt to explore the potential therapeutic effect of inhibiting one of the hits from our analysis, we targeted the glutamine-glutamate-α-ketoglutarate axis by three different strategies, all of which impaired mitochondrial respiration and ATP production and induced apoptosis. Thereby, we report that prednisolone resistance may be accompanied by considerable rewiring of transcriptional and biosynthesis programs. Among other druggable targets that were identified in this study, inhibition of glutamine metabolism presents a potential therapeutic approach in GC-sensitive, but more importantly, in GC-resistant cALL cells. Lastly, these findings may be clinically relevant in the context of relapse-in publicly available datasets, we found gene expression patterns suggesting that in vivo drug resistance is characterised by similar metabolic dysregulation to what we found in our in vitro model.

PRPS2 mutations drive acute lymphoblastic leukemia relapse through influencing PRPS1/2 hexamer stability.

Tumor relapse is the major cause of treatment failure in childhood acute lymphoblastic leukemia (ALL), yet the underlying mechanisms are still elusive. Here, we demonstrate that phosphoribosyl pyrophosphate synthetase 2 (PRPS2) mutations drive ALL relapse through influencing PRPS1/2 hexamer stability. Ultra-deep sequencing was performed to identify PRPS2 mutations in ALL samples. The effects of PRPS2 mutations on cell survival, cell apoptosis, and drug resistance were evaluated. In vitro PRPS2 enzyme activity and ADP/GDP feedback inhibition of PRPS enzyme activity were assessed. Purine metabolites were analyzed by ultra-performance liquid-chromatography tandem mass spectrometry (UPLC-MS/MS). Integrating sequencing data with clinical information, we identified PRPS2 mutations only in relapsed childhood ALL with thiopurine therapy. Functional PRPS2 mutations mediated purine metabolism specifically on thiopurine treatment by influencing PRPS1/2 hexamer stability, leading to reduced nucleotide feedback inhibition of PRPS activity and enhanced thiopurine resistance. The 3-amino acid V103-G104-E105, the key difference between PRPS1 and PRPS2, insertion in PRPS2 caused severe steric clash to the interface of PRPS hexamer, leading to its low enzyme activity. In addition, we demonstrated that PRPS2 P173R increased thiopurine resistance in xenograft models. Our work describes a novel mechanism by which PRPS2 mutants drive childhood ALL relapse and highlights PRPS2 mutations as biomarkers for relapsed childhood ALL.

Relapsed Childhood Acute Lymphoblastic Leukemia: Experience from a Single Tertiary Center in Thailand.

The outcomes of relapsed childhood acute lymphoblastic leukemia (ALL) in developed countries have improved over time as a result of risk-adapted, minimal residual disease-directed therapy, hematopoietic stem cell transplantation, and immunotherapy. There are few studies that have examined survival in relapsed childhood ALL in resource-limited countries. Therefore, this study aimed to assess the prognostic factors and survival outcome of relapsed childhood ALL in a major tertiary center in Southern Thailand. The medical records of patients with ALL aged <15 years between January 2000 and December 2019 were retrospectively reviewed. The Kaplan-Meier method was used to depict the overall survival (OS). A total of 472 patients with ALL were enrolled and relapsed ALL was found in 155 (32.8%) patients. Of these, 131 (84.5%) and 24 (15.5%) had B-cell and T-cell phenotypes, respectively. One hundred thirteen (72.9%) and 42 (27.1%) patients had early and late relapses, respectively. The most common site of relapse was bone marrow in 102 patients (65.8%). One hundred twenty-eight (82.6%) patients received treatment while 27 (17.4%) patients refused treatment. The 5-year OS of all relapsed patients was 11.9%. The 5-year OS among the patients with early relapse was significantly lower than in the patients with late relapse (5.3% vs. 29.1%, respectively, p <0.0001). Site and immunophenotype were not associated with survival of relapsed ALL. The median survival times among the patients who received and refused relapse chemotherapy were 11.8 and 3.1 months, respectively (p <0.0001). The relapse rate accounted for one third of patients with ALL with the 5-year OS of 12%. Early relapse and those who refused treatment were associated with poor survival outcome.

PB1767: OUTCOME OF RELAPSED CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA IN THE SOUTH OF TUNISIA SINGLE CENTER EXPERIENCE

Background: Relapsed acute lymphoblastic leukemia (ALL) has remained challenging to treat in children, with survival rates lagging well behind those observed at initial diagnosis. Although there have been some improvements in outcomes over the past few decades, only ∼50% of children with first relapse of ALL survive long term, and outcomes are much worse with second or later relapses. Aims: In this study we describe clinical characteristics and therapeutics results of first relapse of childhood ALL. Methods: Our study included children under 16 years on first relapse of ALL treated according to the EORTC 58951 protocol between January 2000 and December 2020 in the Hematology department of Hedi Chaker Hospital of Sfax. The marrow relapse was defined as the reapparition in the peripheral smears and/or the increase of blasts (>20%) in bone marrow (BM) after a period of complete remission (CR). A combined relapse included another extra medullary location, neurological (N) or gonadal (G). Relapse treatment was based on the COOPRALL 1997 protocol and since 2008, COPRALL 2007 protocol or the VHR group of the 58951 EORTC protocol. Results: Among the 262 Childs in CR, 81 was relapsed (31%) after a medium follow-up of 127 months. There are 27 females and 54 males. Median age was 7 years and 6 months. Relapse’s frequencies in the groups low risk, average risk 1, average risk 2 and high risk were respectively 4, 39, 18 et 39% of cases. Median of relapse timing was 12 months (2 to 83). Cumulative incidence of relapse at 1, 2 et 3 years after complete remission (CR) were respectively 51, 67 et 84%. Three relapses were occurred after 5 years of CR. Timing of relapse was not correlated with treatment group (p=0.8). The relapse was only medullary in 72% of cases, only neurological in 5% of cases and combined in 19% of cases (M+N= 6%, M+G= 9% et M+N+G= 4%). Second course was delivered to 57% of these patients. Protocol followed was the COOPRALL 1997 in 12 cases (26%), COOPRALL2007 in 28 cases (61%) and EORTC VHR in 6 cases (13%). The second CR (CR2) was get in 55% (44 patients) of treated patients, among them only 6 were allograft (24%). Toxic death before CR had occurred in 19% of treated cases. Among relapsed patients without curative treatment, only one was vivant (neurological relapse). Survivors after first relapse were less than 20%. Summary/Conclusion: In our cohort, relapse’s rate was more than rates reported in other literature series (31% vs 10 to 15%). Relapse’s prognostic factors validated by the literature were the ALL’s phenotype, cytogenetic abnormalities (phi +), timing of relapse, combined relapses (medullary is more serious) and intensity of treatment. Those factors were not been find in our cohort because of the little number of cases. CR2 rates was less than literature (55 vs 80%); the revision of followed relapse’s protocol is to be discussed. As well, the survival of our patients after ALL relapse was less then reported rates (less than 20% vs 35% at three years). This is due to the abstention of treatment in patient in precocious relapse haven’t a sibling familial HLA donor.

Blinatumomab Is Cost-Effective Compared to Standard Chemotherapy for Children with High Risk Relapses of Acute Lymphoblastic Leukemia: A Cost-Effectiveness Analysis Using Population-Based Healthcare Data

▪Introduction: Though cure rates for childhood acute lymphoblastic leukemia (ALL) have improved significantly, outcomes for children with relapsed ALL remain poor. Novel immunotherapies have proved effective in relapsed disease. Blinatumomab, a bispecific T-cell engager targeting CD19, was recently shown to improve disease-free and overall survival among children with high-risk relapsed disease (Brown et al, 2021) prior to proceeding to stem cell transplant. Blinatumomab however also represents a significant cost burden to institutions and healthcare systems. Our objective was thus to determine the cost-effectiveness of blinatumomab vs. standard of care therapy among children with high-risk relapsed ALL.Methods: We used a childhood ALL-specific policy simulation model previously developed using real world clinical, healthcare utilization, and cost population-based data from Ontario, Canada, and restricted it to children with B-lineage ALL who relapsed with 18 months of their original diagnosis to capture a population of patients with high-risk relapse who survived their original month of re-induction chemotherapy. This model was combined with published data from the Children's Oncology Group randomized control study AALL1331 to estimate the long-term survival, healthcare costs and quality adjusted life years associated with two courses of blinatumomab vs. two courses of standard intensive chemotherapy, followed by stem cell transplant. Combining these two sources allowed for the creation of a multi-state survival model and the estimation of the impact of blinatumomab upon relapse and survival. Healthcare costs for the administration of blinatumomab or chemotherapy were sourced from administrative data, provincial formularies, and published sources. Lifetime costs and quality-adjusted life years were calculated while Monte Carlo simulation was used to propagate parameter uncertainty in the cost-effectiveness outcomes. All costs were calculated in Canadian dollars (CAD).Results: Blinatumomab was associated with on average higher life expectancy (43.4 years vs. 36.8 years) and a higher number of quality adjust life years [2.75 QALYs gained, 95 th confidence interval (95CI) -0.44 to 5.99] but higher costs (60,420 CAD, 95CI 26,794 - 94,047) compared to standard intensive chemotherapy. The incremental cost effectiveness ratio (ICER) associated with blinatumomab was 21,970 CAD/QALY gained. However, substantial uncertainty around model parameters resulted in wide confidence intervals around cost-effectiveness outcomes.Discussion: Initial evidence indicates that the use of blinatumomab in high-risk relapse of childhood ALL as compared to standard chemotherapy is associated with a cost-effectiveness ratio that is comparable to or lower than that associated with other interventions recently introduced in pediatric ALL, and represents a cost-effective intervention for this population. However, estimates are based on limited evidence; further research is necessary to better inform several model parameters and thereby decrease uncertainty in cost-effectiveness outcomes. DisclosuresGupta: Jazz Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees.

NUDT15 polymorphism and NT5C2 and PRPS1 mutations influence thiopurine sensitivity in acute lymphoblastic leukaemia cells.

In chemotherapy for childhood acute lymphoblastic leukaemia (ALL), maintenance therapy consisting of oral daily mercaptopurine and weekly methotrexate is important. NUDT15 variant genotype is reportedly highly associated with severe myelosuppression during maintenance therapy, particularly in Asian and Hispanic populations. It has also been demonstrated that acquired somatic mutations of the NT5C2 and PRPS1 genes, which are involved in thiopurine metabolism, are detectable in a portion of relapsed childhood ALL. To directly confirm the significance of the NUDT15 variant genotype and NT5C2 and PRPS1 mutations in thiopurine sensitivity of leukaemia cells in the intrinsic genes, we investigated 84 B‐cell precursor‐ALL (BCP‐ALL) cell lines. Three and 14 cell lines had homozygous and heterozygous variant diplotypes of the NUDT15 gene, respectively, while 4 and 2 cell lines that were exclusively established from the samples at relapse had the NT5C2 and PRPS1 mutations, respectively. Both NUDT15 variant genotype and NT5C2 and PRPS1 mutations were significantly associated with DNA‐incorporated thioguanine levels after exposure to thioguanine at therapeutic concentration. Considering the continuous exposure during the maintenance therapy, we evaluated in vitro mercaptopurine sensitivity after 7‐day exposure. Mercaptopurine concentrations lethal to 50% of the leukaemia cells were comparable to therapeutic serum concentration of mercaptopurine. Both NUDT15 variant genotype and NT5C2 and PRPS1 mutations were significantly associated with mercaptopurine sensitivity in 83 BCP‐ALL and 23 T‐ALL cell lines. The present study provides direct evidence to support the general principle showing that both inherited genotype and somatically acquired mutation are crucially implicated in the drug sensitivity of leukaemia cells.

Identification of a Seven-lncRNA-mRNA Signature for Recurrence and Prognostic Prediction in Relapsed Acute Lymphoblastic Leukemia Based on WGCNA and LASSO Analyses.

Abnormal expressions of long noncoding RNAs (lncRNAs) and protein-encoding messenger RNAs (mRNAs) are important for the development of childhood acute lymphoblastic leukemia (ALL). This study developed an lncRNA-mRNA integrated classifier for the prediction of recurrence and prognosis in relapsed childhood ALL by using several transcriptome data. Weighted gene coexpression network analysis revealed that green, turquoise, yellow, and brown modules were preserved across the TARGET, GSE60926, GSE28460, and GSE17703 datasets, and they were associated with clinical relapse and death status. A total of 184 genes in these four modules were differentially expressed between recurrence and nonrecurrence samples. Least absolute shrinkage and selection operator analysis showed that seven genes constructed a prognostic signature (including one lncRNA: LINC00652 and six mRNAs: INSL3, NIPAL2, REN, RIMS2, RPRM, and SNAP91). Kaplan-Meier curve analysis observed that patients in the high-risk group had a significantly shorter overall survival than those of the low-risk group. Receiver operating characteristic curve analysis demonstrated that this signature had high accuracy in predicting the 5-year overall survival and recurrence outcomes, respectively. LINC00652 may function by coexpressing with the above prognostic genes (INSL3, SNAP91, and REN) and lipid metabolism-related genes (MIA2, APOA1). Accordingly, this lncRNA-mRNA-based classifier may be clinically useful to predict the recurrence and prognosis for childhood ALL. These genes represent new targets to explain the mechanisms for ALL.

The Effects of Three ABCG2 Polymorphisms on Outcome of Central Nervous System Relapses in Iranian Children With Acute Lymphoblastic Leukemia Receiving High Dose Methotrexate

Methotrexate (MTX) is the main drug for the treatment of childhood acute lymphoblastic leukemia (ALL). ABCG2 pump is the main transporter of MTX on BBB. Our aim was to investigate the possible relationship between three polymorphisms of the ABCG2 gene, and isolated CNS relapses in Iranian children with ALL receiving high dose MTX. Genotyping of three polymorphisms of the ABCG2 gene, including G34A, C376T, and C421A, was performed by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for 56 patients. A high frequency of C376T CT genotype was observed among the patients. There was no significant association between C376T and C421 and isolated CNS relapse (P&gt;0.05). C376T and C421A polymorphisms are not associated with isolated CNS relapse in childhood ALL.

FPGS relapse-specific mutations in relapsed childhood acute lymphoblastic leukemia

Although the cure rate for childhood acute lymphoblastic leukemia (ALL) has exceeded 80% with contemporary therapy, relapsed ALL remains a leading cause of cancer-related death in children. Relapse-specific mutations can be identified by comprehensive genome sequencing and might have clinical significance. Applying whole-exome sequencing to eight triplicate samples, we identified in one patient relapse-specific mutations in the folylpolyglutamate synthetase (FPGS) gene, whose product catalyzes the addition of multiple glutamate residues (polyglutamation) to methotrexate upon their entry into the cells. To determine the prevalence of mutations of the FPGS mutations, and those of two important genes in the thiopurine pathway, NT5C2 and PRPS1, we studied 299 diagnostic and 73 relapsed samples in 372 patients. Three more FPGS mutants were identified in two patients, NT5C2 mutations in six patients, and PRPS1 mutants in two patients. One patient had both NT5C2 and PRPS1 mutants. None of these alterations were detected at diagnosis with a sequencing depth of 1000X, suggesting that treatment pressure led to increased prevalence of mutations during therapy. Functional characterization of the FPGS mutants showed that they directly resulted in decreased enzymatic activity, leading to significant reduction in methotrexate polyglutamation, and therefore likely contributed to drug resistance and relapse in these cases. Thus, besides genomic alterations in thiopurine metabolizing enzymes, the relapse-specific mutations of FPGS represent another critical mechanism of acquired antimetabolite drug resistance in relapsed childhood ALL.

Mutational landscape and patterns of clonal evolution in relapsed pediatric acute lymphoblastic leukemia.

Relapse of acute lymphoblastic leukemia (ALL) remains a leading cause of childhood death. Prior studies have shown clonal mutations at relapse often arise from relapse-fated subclones that exist at diagnosis. However, the genomic landscape, evolutionary trajectories and mutational mechanisms driving relapse are incompletely understood. In an analysis of 92 cases of relapsed childhood ALL, incorporating multimodal DNA and RNA sequencing, deep digital mutational tracking and xenografting to formally define clonal structure, we identify 50 significant targets of mutation with distinct patterns of mutational acquisition or enrichment. CREBBP, NOTCH1, and Ras signaling mutations rose from diagnosis subclones, whereas variants in NCOR2, USH2A and NT5C2 were exclusively observed at relapse. Evolutionary modeling and xenografting demonstrated that relapse-fated clones were minor (50%), major (27%) or multiclonal (18%) at diagnosis. Putative second leukemias, including those with lineage shift, were shown to most commonly represent relapse from an ancestral clone rather than a truly independent second primary leukemia. A subset of leukemias prone to repeated relapse exhibited hypermutation driven by at least three distinct mutational processes, resulting in heightened neoepitope burden and potential vulnerability to immunotherapy. Finally, relapse-driving sequence mutations were detected prior to relapse using deep digital PCR at levels comparable to orthogonal approaches to monitor levels of measurable residual disease. These results provide a genomic framework to anticipate and circumvent relapse by earlier detection and targeting of relapse-fated clones.

TP53 alterations in relapsed childhood acute lymphoblastic leukemia.

TP53 alterations are frequent relapse‐acquired mutations in childhood acute lymphoblastic leukemia (ALL). The present study evaluated the clinical significance of relapsed childhood ALL in Taiwan. Diagnostic and/or relapsed bone marrow or peripheral blood was obtained from 111 children with relapsed ALL who were initially treated by using Taiwan Pediatric Oncology Group (TPOG) ALL protocols from January 1997 to May 2018. Mutations were detected by PCR and sequencing, as well as by multiplex ligation‐dependent probe amplification to detect copy number alterations. Copy number and/or sequence alterations of TP53 were detected in 29% (28 of 98) and in 46% (6 of 13) of patients with relapsed B‐cell and T‐cell ALL, respectively. This incidence was much higher than that in several similar studies conducted in Caucasian populations. Seventy percent of all TP53 alterations were gained at relapse in 67 matched samples by back‐tracking matched diagnostic samples. TP53 alterations were associated with lower 5‐year event‐free survival (EFS) and overall survival (OS) rates (P = .013 and P = .0002, respectively). Multivariate analysis confirmed the prognostic significance of TP53 alterations. Forty‐five patients received hematopoietic stem‐cell transplantations post‐relapse. Patients with TP53 alterations (14/45) had inferior 5‐year EFS and OS than patients without TP53 alterations after transplantation (P = .002 and P = .001, respectively). The significance of these TP53 alterations for patients who received transplantations was confirmed by multivariate analysis. In conclusion, TP53 alterations were enriched and useful as prognostic markers in relapsed childhood ALL.

SP-38 - Novel therapy for relapsed childhood acute lymphoblastic leukemia

SP-38 - Novel therapy for relapsed childhood acute lymphoblastic leukemia

Relapsed childhood acute lymphoblastic leukemia: Current situation in China; a multicenter observational study

BackgroundRelapse is the major reason for death in pediatric acute lymphoblastic leukemia (ALL) patients. It may be worse because of refusing re-treatment in developing countries. So, we investigated the influencing factors for re-treatment decision made by parents and survival data of patients with relapsed pediatric ALL in China. MethodsClinical characteristics, choice of re-treatment, survival status at the last follow-up time were collected, survival data and prognostic factors were analyzed. ResultsAmong 390 relapsed cases which were collected from five centers in China, 230 (59.0%) were re-treated. Fifty-eight were alive, 249 had died, and 83 were lost to follow up (median follow up 73.0 months). Receiving retreatment was not associated with gender, initial immunophenotype, or era of relapse, but was associated with age, initial risk group, and recurrence time and site. The 5-year OS rate of patients who received re-treatment or not was 35.5% and 4.5%, respectively. In re-treatment group, the survival analysis showed that initial risk group, site of relapse, time from diagnosis to relapse, and whether the patient achieved CR2 were closely related to 5-year OS, while WBC count (50 × 109/L), and immunophenotype at diagnosis were not. Fortyeight cases received transplantation and 20 (41.7%) survived, while only 36 (19.8%) of 182 patients who did survived. ConclusionsThe estimated 5-year OS rate was only 24.4% ± 2.5% for all 390 relapsed cases in China. Of them, 59% patients, 230 cases, were re-treated with 5-year OS 35.5%. Patients who were indicated and performed for HSCT had a 5-year OS of 52.2%.

"What's in a name?" CAR-T Gene Therapy.

With all due respect to Shakespeare, the impact of nomenclature should not be underestimated. Words are both effective and affective. They send a signal to the listener but may have an even more pronounced effect on the mind of the speaker. During a four-year stint I finished in 2010 on the National Institutes of Health Recombinant DNA Advisory Committee, the committee was repeatedly reminded that we should not speak of “gene therapy.” “Gene transfer research” (the preferred terminology) was resurgent in those years as the science made progress in safety and efficacy, and some time had passed since the tragic death of Jesse Gelsinger as a direct result of his participation in such a study. But by no means was the language of treatment considered appropriate—at least according to the powers that be at NIH. I suspect that investigators and patients who were considering enrollment on gene-transfer clinical trials might have had a different opinion. The former were attempting to develop treatments for a variety of diseases, and the latter wanted to benefit from the clinical science. Fast forward to August of 2017, when the Food and Drug Administration approved chimeric antigen receptor T cell (CAR-T) therapy for relapsed childhood acute lymphoblastic leukemia (ALL). This ex vivo genetic manipulation of a patient's own T-lymphocytes marks a major success in translational research, a landmark in cancer history, and a sign of good things to come in personalized medicine. Unfortunately, since CAR-T infusion is often associated with a life-threatening cytokine release syndrome, the approval was limited to a small number of centers with the expertise to manage this toxicity. Despite the risk, this is the most dazzling clinical science to come along in the nearly thirty years since I started my fellowship in pediatric oncology. And yes, with the imprimatur of the FDA, it is now acceptable in this case to use the term “gene therapy.” The question of what to name this activity reflects a longstanding debate in research ethics. Is it ever correct to talk about “therapeutic research?” What are the risks of promoting false hopes? During the era in which many aspire to build a learning health care system, perhaps one size does not fit all. High-risk, high-reward research like that which led to the development of CAR-T therapy seems to me to be categorically different from low-risk population-based research that leverages the electronic medical record and the power of big data. Thoughtful attention to such distinctions and to key transition points like FDA approval is the foundation for thinking about research, innovation, progress, and treatment in biomedicine. So, as long as there is a commitment to quality informed consent processes, count me in as comfortable talking about “therapeutic research.” The story of CAR-T doesn't end here, though. The cost is slated to be $475,000 per treatment course, with the expectation that one infusion will be sufficient. The chutzpah of charging a patient that much for getting an infusion of his or her own cells (albeit transformed) is considerable. Other recent breakthrough drugs for cystic fibrosis and hepatitis C have also been triggers for sticker shock. What are the justifications for such pricing, and how, if at all, might we think about limiting the cost? With a commitment to free-market approaches to drug development, we may be moving inexorably toward creating unobtainable cures. Progress against cancer happens in fits and starts. CAR-T represents a double-edged sword, a seemingly miraculous start coupled with a cautionary tale of clinical risk and cost. Having cared for many children who have died of ALL and having also seen how the late effects of our best treatments can impair quality of life, I pray that in the coming years we can maximize the benefits, minimize the cost and the harm, and help cancer patients survive and thrive. I would call that success.

CREBBP knockdown enhances RAS/RAF/MEK/ERK signaling in Ras pathway mutated acute lymphoblastic leukemia but does not modulate chemotherapeutic response.

Relapsed acute lymphoblastic leukemia is the most common cause of cancer-related mortality in young people and new therapeutic strategies are needed to improve outcome. Recent studies have shown that heterozygous inactivating mutations in the histone acetyl transferase, CREBBP, are particularly frequent in relapsed childhood acute lymphoblastic leukemia and associated with a hyperdiploid karyotype and KRAS mutations. To study the functional impact of CREBBP haploinsufficiency in acute lymphoblastic leukemia, RNA interference was used to knock down expression of CREBBP in acute lymphoblastic leukemia cell lines and various primagraft acute lymphoblastic leukemia cells. We demonstrate that attenuation of CREBBP results in reduced acetylation of histone 3 lysine 18, but has no significant impact on cAMP-dependent target gene expression. Impaired induction of glucocorticoid receptor targets was only seen in 1 of 4 CREBBP knockdown models, and there was no significant difference in glucocorticoid-induced apoptosis, sensitivity to other acute lymphoblastic leukemia chemotherapeutics or histone deacetylase inhibitors. Importantly, we show that CREBBP directly acetylates KRAS and that CREBBP knockdown enhances signaling of the RAS/RAF/MEK/ERK pathway in Ras pathway mutated acute lymphoblastic leukemia cells, which are still sensitive to MEK inhibitors. Thus, CREBBP mutations might assist in enhancing oncogenic RAS signaling in acute lymphoblastic leukemia but do not alter response to MEK inhibitors.

Mutational Landscape of Pediatric Acute Lymphoblastic Leukemia Relapsing after Allogeneic Stem Cell Transplantation

The prognosis of children with acute lymphoblastic leukemia (ALL) relapsing following allogeneic hematopoietic stem cell transplantation (allo-SCT) is still dismal. Within the framework of the ALL-REZ BFM 2002/ALL-SZT BFM 2003 trials, we performed whole-exome sequencing (WES) of ten patients relapsing after allo-SCT with the aim to thoroughly characterize the spectrum of acquired mutations and to identify potentially druggable targets, thus laying the ground for future personalized treatment strategies.Patients’ age at initial diagnosis ranged from <12 months to 10 years. Two patients were diagnosed with T-ALL and 8 with pre-B-ALL. Time from initial disease to relapse ranged from 13 to 40 months. Patients underwent allo-SCT 3 to 6 months after diagnosis of relapse, donors were either HLA-matched unrelated volunteers (n=6), HLA-identical siblings (n=1) or HLA-haploidentical family donors (n=1). Time from SCT to relapse ranged from 3 to 21 months. Treatment following the post allo-SCT relapse varied considerably (from palliative care to a second allo-SCT). Only 2/10 patients are still alive, with one recently having been diagnosed with subsequent relapse following a second SCT.To investigate the mutational landscape of relapsed ALL following high-dose chemotherapeutic and immunologic attack both provided by an allo-SCT, five samples per patient were analyzed: initial leukemia (INIT), remission (REMI) after front-line chemotherapy representing patient germline, first relapse (RLPS), full donor-chimeric remission post allo-SCT (TREMI) representing donor germline, and relapse post allo-SCT (TRLPS). For comparative analyses, we defined the following three "oncogenomes" (OGs): OG1 (initial leukemia, SNVs in INIT minus REMI)), OG2 (first relapse, SNVs in RLPS minus REMI), OG3 (post allo-SCT relapse, SNVs in TRPLS minus REMI or TREMI).Median numbers of leukemia-specific SNVs in OG1-3 were 8.5, 34 and 37.5. We detected a median of 0.18 mutations per megabase (MB) in OG1, 0.67 mutations/MB in OG2 and 0.74 mutations/MB in OG3 (p= 0.005 for OG2/3 vs. OG1 [Wilcoxon test]).Although the mutational spectrum was highly diverse between individual patients and within the oncogenomes, we also identified several recurrent alterations: in OG1, five genes had recurrent SNVs (IGSF3, TTN, NOTCH1, CTBP2, NRAS). Seven genes were recurrently affected in three OG2s, including IKZF1, NOTCH1, NRAS, NT5C2. In OG3 we detected eleven recurrently mutated genes in three patients, among which were NRAS, FLT4, and TP53.Notably, TP53 was mutated in 5/10 patients. One patient carried a TP53 germline mutation, one patient had one unique SNV each in OG2 as well as OG3 and three patients had TP53 mutations only present in their OG3s. All but one SNV (resulting in a premature stop codon) were non-synonymously coding and were predicted to be deleterious for protein function.Of particular note, leukemic blasts showed profound plasticity concerning the mutational status of the nucleoside exporter NT5C2, mutations of which were previously shown to drive chemotherapy resistance in relapsed childhood ALL. While NT5C2 alterations were completely absent in OG1, 4 SNVs were detected in the OG2s of 3/10 patients. However, these NT5C2 mutations disappeared again in the OG3s once selection pressure of maintenance chemotherapy employing nucleoside analogues had been withdrawn.To identify novel treatment options in the desperate clinical scenario of post allo-SCT relapse, we searched OG3 for genetic lesions in genes known to be either targeted directly (a certain gene) or indirectly (a certain pathway) by currently approved therapeutic agents. To our surprise, nine out of ten patients exhibited such SNVs, for which additional targeted therapies are already available. Those therapeutic agents comprise small molecules inhibitors as well as antibodies such as Dasatinib, Erlotinib, Ibrutinib, Pazopanib, Tocilizumab, and Trastuzumab.Conclusion:Our comprehensive genetic analysis of ten children with ALL relapsing post allo-SCT revealed profound leukemic cell plasticity (e.g. loss of acquired NT5C2 mutations) as well as identified several recurrent genetic alterations (e.g. NOTCH1, NRAS, IKZF1) including TP53 (5/10 patients). Most importantly, we identified alterations in genes amenable to targeted treatment approaches in 9/10 patients thus potentially opening new therapeutic avenues. DisclosuresBader:Novartis: Consultancy, Honoraria; Servier: Consultancy, Honoraria; Neovii Biotech: Research Funding; Riemser: Research Funding; Medac: Consultancy, Research Funding. Peters:Novartis: Consultancy; Jazz: Speakers Bureau; Amgen: Consultancy; Pfizer: Consultancy; Medac: Consultancy.