Adult ALL Research Articles

Transplant in Adult ALL: Who and When

Transplant in Adult ALL: Who and When

Significance of Measurable Residual Disease in Adult Philadelphia Chromosome-Positive ALL: A GRAAPH-2014 Study.

BCR::ABL1 quantification is widely regarded as the standard for monitoring measurable residual disease (MRD) in Philadelphia chromosome-positive (Ph+) ALL. However, recent evidence of BCR::ABL1 multilineage involvement questions the significance of BCR::ABL1 MRD. We aimed to define the prognostic role of MRD as assessed by BCR::ABL1 or lymphoid-specific immunoglobulin/T-cell receptor (IG/TR) gene markers. We conducted BCR::ABL1 and IG/TR quantification after each treatment cycle in 264 patients treated in the GRAAPH-2014 trial, which used four cycles of reduced-intensity chemotherapy with nilotinib, followed by hematopoietic stem-cell transplantation (HSCT). Comparing BCR::ABL1 and IG/TR MRD revealed residual BCR::ABL1-positive non-ALL cells in 98 (43%) of 228 patients, defining multilineage Ph+ ALL. Despite poorer BCR::ABL1 responses, patients with multilineage Ph+ ALL had similar disease-free survival (DFS; hazard ratio [HR], 0.83 [95% CI, 0.49 to 1.41]; P = .50). Although BCR::ABL1 response failed to predict outcomes, IG/TR positivity (≥0.01%) was strongly associated with lower DFS (after cycle 2, HR, 2.49 [95% CI, 1.40 to 4.40]; P = .002; after cycle 4, HR, 4.13 [95% CI, 1.82 to 9.38]; P = .001). In multivariable analysis, both IG/TR positivity after cycle 2 and initial WBC count ≥30 × 109/L predicted poorer DFS, enabling to define a high-risk group having a 4-year DFS of 56.5% compared with 87.6% (HR, 3.72 [95% CI, 1.93 to 7.15]; P < .001). Moreover, allogeneic HSCT significantly improved DFS in the high-risk group (HR, 0.33 [95% CI, 0.18 to 0.60]; P < .001), whereas the standard-risk group had favorable outcomes regardless of allogeneic HSCT. Our findings challenge the significance of BCR::ABL1 monitoring in adult Ph+ ALL and demonstrate the prognostic role of IG/TR MRD. This study provides a framework for using MRD to guide treatment strategies in adults with Ph+ ALL.

A Multicentre, Randomized Trial in Adults with de novo Philadelphia Chromosome-Positive Acute Lymphoblastic Leukaemia to Assess the Efficacy of Ponatinib versus Imatinib in Combination with Low-Intensity Chemotherapy, to Compare End of Therapy with Indication for Stem Cell Transplantation versus Tyrosine Kinase Inhibitor, Blinatumomab, and Chemotherapy in Optimal Responders, and to Evaluate Blinatumomab in Suboptimal Responders (GMALL-EVOLVE)

Introduction: Philadelphia chromosome-positive acute lymphoblastic leukaemia (Ph+ALL) is treated as standard of care (SoC) by imatinib-based treatment combined with induction and consolidation chemotherapy followed by allogeneic stem cell transplantation (SCT) in first remission. The German Multicenter ALL Study Group for Adult ALL (GMALL) reports about a trial to evaluate the impact of ponatinib-based therapy, blinatumomab treatment for suboptimal responders, and the possibility of omission of SoC Allo SCT in optimal responders entitled GMALL-EVOLVE. Methods: Herein, imatinib is randomized versus ponatinib as frontline treatment combined with chemotherapy, optimal responders also get randomized between SCT and chemo-immunotherapy, and suboptimal responders receive immunotherapy before SCT. The trial is registered under the EudraCT number 2022-000760-21. Conclusion: This trial will answer several major questions in the treatment of Ph+ALL.

Impact of the COVID-19 pandemic on utilization and cost for care of pediatric and young adult ALL

ObjectiveAcute lymphoblastic leukemia (ALL) is the most common childhood malignancy and among the most common malignancies in young adults and requires a unique pattern of healthcare utilization including an acute/emergent presentation and an intensive initial 8 months of therapy followed by two years of outpatient treatment. The COVID-19 pandemic caused massive global disruptions in healthcare use and delivery. This report aims to examine the effects of the COVID-19 pandemic on the presentation, diagnosis and continued management of childhood and young adult ALL in regard to utilization and cost of care among commercially insured individuals in the United States.ResultsUtilizing a commercial insurance claims database, 529 pediatric and young adult patients were identified who were diagnosed with ALL between January 2016 and March 2021. New diagnoses were evaluated by era and demographics. Utilization was measured by COVID-related era as number of inpatient and outpatient encounters, inpatient days, and cumulative cost during the initial 8 months of therapy. None of these cost or utilization factors changed significantly during or shortly after the pandemic. These findings reinforce that the necessary care for pediatric and young adult ALL was unwavering despite the massive shifts in the healthcare system caused by the COVID-19 pandemic. This provides a valuable benchmark as we further examine the factors that influence the pandemic’s impact on health equity and access to care, especially in vulnerable pediatric and young adult populations. This is the first investigation of the effect of the COVID-19 pandemic on utilization and cost of care in pediatric and young adult cancer.

Real world outcome of B ALL with t (1; 19) (q23; p13)/TCF3::PBX1 in adolescents and adults treated with intensive regimes

Significant heterogeneity has been reported in outcome of Acute lymphoblastic leukemia with t(1;19)(q23;p13)/TCF3::PBX1 in adolescents and adults leading to a lack of consensus on precise risk stratification. We evaluated clinical outcome of 17 adult ALL cases (≥15 years) with this genotype treated on intensive regimes.13/17 received COG0232 and 4/17 cases received UK-ALL protocol. All achieved CR (100%) with above treatment. End of induction MRD was evaluated in 14/17 cases of which 11 (78.5%) achieved MRD negativity. Total nine patients relapsed (7 marrows, 2 CNS). Overall survival at 2 years was 53.3%. The 2 year estimated PFS was 42.9%. The 2 years CIR was 54.2%. Adults with this genotype perform poorly despite early favorable response. Incorporation of novel immunotherapies and prompt HSCT should be strongly considered with this genotype. Targeted NGS panels for additional genetic aberrations can further help in risk stratifying and guiding therapy for this genotype.

Abstract 5956: FLT3 alterations and inhibition in triple negative B-cell adult acute lymphoblastic leukemia patients

Abstract Backgroud: The prognosis of B cell ALL patients (pts) relapsing after Hematopoietic Transplant is poor and new drugs are needed. In acute myeloid leukemia, FLT3-inhibitors (FLT3i) are showing promising results in FLT3-mutated (mut) cases. Their potential use in FLT3-mut B-ALL has been poorly investigated. Aim: To assess if FLT3 could be a valid targetable marker in B-ALL. Methods: We sequenced with a capture based large RNA-seq panel (1385genes) 183 adult B-ALL [n=52 Pos for Ph or t(4;11) or t(1;19) and n=131 Triple-Negative (TN) pts that are negative for Ph (n=43), t(4;11) and t(1;19) translocations] and 15 donors. To confirm FLT3 muts, 13 available samples were further sequenced with Extended Myeloid Solution (98 genes; SOPHiA) where FLT3 gene is fully covered. In vitro studies using 5 FLT3i [Gilteritinib (Gil), Midostaurin, Crenolanib, Sorafenib and Quizartibin] and Venetoclax (Ven) were conducted on pt primary cells and on TN and other B-ALL cell lines (MUTZ5, MHH-CALL4 Ph-like; NALM6;RS4;11, 697, REH) and 2 AML cell lines (OCI-AML3 FLT3;MV-4-11 FLT3-ITD). Results: We found 15 FLT3 mut in 14/131 TN B-ALL and in 1/43 Ph+ cases: 43.8% were TKD mut, 3 ITD mut, 3 splicing site mut,1 N-terminal, 1 juxta-membrane domain and 1 Ig-like site (Fig. 1A). After pathogenicity interpretation we excluded 3 variants and one mutation was not confirmed at DNA level. Overall 7.6% of TN were mut (Ph+ 2.3%). 81.8% of FLT3mut pts potentially druggable with FLT3i, mainly TKD (63.3%). Targeted NGS revealed the co-occurrence with other mutations (range 2-15), and in particular with CSMD1 (n=4), KDM6A, KMT2D and CREBBP (n=3) alterations. Moreover, 11/13 samples carried copy number alteration (mean 12), mostly amplifications. CDKN2A &amp; IKZF1 were the most frequently altered genes (n=5;4 CN-loss). FLT3 expression was increased in 11/15 cases compared to donors and wt pts. As reported, the expression was higher in ZNF384 or MLLr B-ALL subtypes.To evaluate the effect of FLT3is in ALL, we treated ex vivo primary leukemic cells of 6 adult ALL patients (FLT3-mut n=4; FLT3-wt n=2) with increasing concentrations of 5 FLT3i for 24, 48 and 72h. We observed a trend towards greater response of FLT3-mut ALL cells compared to wt ones. The average IC50 values in response to the five FLT3i was 1.6 uM and 24.5 uM after 72h of treatment in FLT3-mut and FLT3-wt B-ALL, respectively (Fig. 1B). We than evaluated the sensitivity to the 5 Inhs in all cell lines. Interestingly MHH-CALL4 wt were more sensitive to Gil compared to OCI AML3 wt, and showed the same effect compare to MV-4-11 AML mut (F.1C). Given the effect of Gil in single agent, we combined it with Ven in wt B-ALL cells at increasing concentrations. Interestingly We noticed a strong additive effect with the higher Gil concentration and all Ven doses(F.1D). Conclusions: FLT3 alterations identify a novel subgroup of TN B-ALL with therapeutic potential also in combination regimens. Ricerca Corrente Italian MoH L3P1946. Citation Format: Anna Ferrari, Andrea Ghelli Luserna di Rorà, Federico Lo Schiavo, Eugenio Fonzi, Giorgia Capirossi, Chiara Salvesi, Maria Teresa Bochicchio, Lorenzo Ledda, Chiara Servilli, Matteo Paganelli, Giovanni Marconi, Cristina Papayannidis, Chiara Sartor, Michela Rondoni, Anna Maria Mianulli, Barbara Giannini, Crescenza Pasciolla, Fabio Giglio, Sara Galimberti, Monica Fumagalli, Prassede Salutari, Valentina Gaidano, Giovanni Martinelli, Giorgia Simonetti. FLT3 alterations and inhibition in triple negative B-cell adult acute lymphoblastic leukemia patients [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 5956.

Abstract 3910: The Atlas of Blood Cancer Genomes: A resource for therapeutic and biomarker development

Abstract Developing a novel cancer therapy is an expensive, time-consuming, high-risk endeavor that involves identifying a molecular target as well as target indications. This process could be accelerated by a comprehensive interrogation of driver variants and gene expression profiles across cancer types. The Atlas of Blood Cancer Genomes (ABCG) project was initiated to elucidate the molecular basis of all leukemias and lymphomas, building on advances in genomic technologies, our capabilities for data analysis, and economies of scale. The ABCG project includes collaborators from 25 institutions worldwide who contributed samples from 10,512 patients comprising every type of blood cancer in the World Health Organization classifications. All cases were de-identified and their associated pathology and clinical information entered into a purpose-built web-based system. All cases underwent pathology and clinical data review by experienced hematopathologists and oncologists. Samples were subjected to whole exome DNA and RNA sequencing. We examined three classes of therapeutic targets with selected examples of application: 1. Surface markers: Surface markers are targets of many approved and experimental therapies in blood cancers (CAR-T cells, monoclonal and bispecific antibodies). We found that CD22, which has been evaluated as a target in diffuse large B-cell lymphoma (DLBCL), is also highly expressed in follicular lymphoma, adult ALL, mantle cell lymphoma, and high-grade B cell lymphoma, all areas of clinical need. We can evaluate the expression of virtually any marker or combination of markers across all blood cancers while noting areas of greatest clinical need within and across diseases, providing the basis to reclassify diseases by therapeutic target. 2. Genetic targets: Our work assesses the entire spectrum of genetic alterations including mutations and fusions. We found targetable alterations including BCR-ABL1 fusions, EZH2 Y641, IDH2 R140Q and BRAF V600E mutations in multiple cancers, albeit at low frequency, pointing to potential new indications for existing drugs in subsets of rare diseases. 3. Complex targets (immune or expression signatures or combinations of gene variants): Past work defined DLBCL immune and other signatures associated with response to avadomide, a novel cereblon inhibitor. We found that these signatures are also highly expressed in large subsets of peripheral T-cell lymphomas and acute myeloid leukemia, both areas of major unmet clinical need. Thus, we can interrogate immune and other signatures across the spectrum of cancers to uncover potential biomarkers of response. The ABCG project will enable the comprehensive study of genomic and clinicopathological features of all blood cancers. We anticipate that our data, approaches and results will provide a lasting resource for molecular classification and therapeutic development in all leukemias and lymphomas. Citation Format: Jennifer Shingleton, Raju Pillai, Sarah Ondrejka, Govind Bhagat, Amy Chadburn, Matthew McKinney, Jean Koff, Dina Soliman, Magdalena Czader, Abner Louissaint, Shaoying Li, Choon Kiat Ong, Amir Behdad, Andrew Evens, Yaso Natkunam, Mette Pedersen, Sirpa Leppa, Eric Tse, Jennifer Chapman, Catalina Amador-Ortiz, Yuri Fedoriw, Andrew Evans, Jiong Yan, Mina Xu, Kikkeri Naresh, Clay Parker, David Hsu, Sandeep Dave. The Atlas of Blood Cancer Genomes: A resource for therapeutic and biomarker development [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 3910.

Abstract 3206: Tumor-targeted nanoparticles for cancer therapy

Abstract Introduction: Untargeted cancer therapy is often toxic, limiting the maximum tolerated dose (MTD) and thus potential cure. Targeted therapy with antibody-drug conjugates (ADCs) has demonstrated greater efficacy, as with those directed at HER2 for triple negative breast cancer and other HER2 expressing tumors. However, ADC drug-antibody ratio (DAR) is limited, typically 3 to 1. In contrast, antibody-targeted nanoparticles deliver thousands of drug molecules per nanoparticle, with potentially greater anti-tumor effect. Experimental Procedures: Antibody targeted nanoparticles (TNP) were created that preferentially deliver cytotoxic molecules to tumor cells. TNPs were created with various antibodies and payloads and evaluated for their efficacy against 12 different adult and pediatric malignancies (see below), using tumor xenografts in NOG-SCID mice. Survival of control, free drug, and TNP treated animals was compared. MED was also determined. Non-tumor bearing animals treated at the same dose and schedule were evaluated for evidence of toxicity. PK/PD metrics were obtained and drug delivery to normal and tumor tissue was evaluated microscopically and by mass spectroscopy. Data: Plasma TNP drug levels persist for 48 hours, compared to free drug which is undetectable by 2 hours. Microscopically, nanoparticles accumulate in tumor tumor tissue but not normal tissue. Drug levels are typically 15 times higher in tumor tissue than normal tissue by quantitative mass spectroscopy. The minimum effective dose (MED) is reduced 25-fold for Ewing sarcoma, from 50 mg/kg to 2 mg/kg. TNPs also cross the blood-brain barrier (BBB), and were used to treat orthotopic GBM PDX xenografts, prolonging animal survival 3-fold over control or free drug treated animals. Ewing sarcoma, osteosarcoma, triple negative breast cancer, GBM, hepatoma, and adult ALL, bearing animals all showed complete tumor ablation and 100% survival when treated biweekly at 10mg/kg. Pancreatic, prostate, and ovarian Ca showed 100% survival in TNP treated mice after all control animals had died but remained tumor bearing with slow but progressive tumor growth when treated identically. Lung and colorectal Ca and melanoma bearing animals treated similarly with TNPs showed 50-80% survival at the time of control animal death. No animal in any study showed evidence of off-target toxicity. Conclusions: Tumor antigen targeted, cytotoxic molecule containing nanoparticles have been created that show broad utility in the treatment of at least a dozen different adult and pediatric cancers. Six of 12 types showed complete tumor ablation when treated at 10mg/kg biweekly injections, and the remainder showed enhanced anti-tumor effect and survival compared to free drug treatment. We conclude that use of tumor antigen targeted, small molecule cytotoxic containing nanoparticles results in dramatically enhanced tumor response and in many case, complete tumor ablation where the same free drug fails to do so. Citation Format: HyungGyoo Kang, Bryon Upton, Jon Nagy, Timothy Triche. Tumor-targeted nanoparticles for cancer therapy [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 3206.

Efficacy of allogeneic hematopoietic stem cell transplantation based on a total body irradiation conditioning treatment regimen for adult acute lymphocytic leukemia

Objective: To analyze the efficacy of allo-HSCT with total body irradiation (TBI) and chemotherapy alone in the treatment of adult ALL and to explore the factors affecting prognosis. Methods: The clinical data of 95 adult patients with ALL who underwent allo-HSCT from January 2015 to August 2022 were included. According to the conditioning regimen, the patients were divided into two groups: the TBI plus cyclophosphamide (TBI/Cy) group (n=53) and the busulfan plus cyclophosphamide (Bu/Cy) group (n=42). Hematopoietic reconstitution after transplantation, GVHD, transplantation-related complications, relapse rate (RR), non-relapse mortality (NRM), OS, and LFS were compared, and the factors related to prognosis were analyzed. Results: The median time of neutrophil engraftment was 14 (10-25) days in the TBI/Cy group and 14 (10-24) days in the Bu/Cy group (P=0.106). The median time of megakaryocyte engraftment was 17 (10-42) days in the TBI/Cy group and 19 (11-42) days in the Bu/Cy group (P=0.488). The incidence of grade Ⅱ-Ⅳ acute GVHD (aGVHD) in the TBI/Cy and Bu/Cy groups was 41.5% and 35.7%, respectively (P=0.565). The incidence of grade Ⅲ-Ⅳ aGVHD in these two groups was 24.5% and 4.8%, respectively (P=0.009). The incidence of severe chronic GVHD in the two groups was 16.7% and 13.5%, respectively (P=0.689). The incidence of cytomegalovirus infection, Epstein-Barr virus infection, severe infection, and hemorrhagic cystitis in the two groups was 41.5% and 35.7% (P=0.565), 34.0% and 35.7% (P=0.859), 43.4% and 33.3% (P=0.318), and 20.8% and 50.0% (P=0.003), respectively. The median follow-up time was 37.1 months and 53.3 months in the TBI/Cy and Bu/Cy groups, respectively. The 2-year cumulative RR was 17.0% in the TBI/Cy group and 42.9% in the Bu/Cy group (P=0.017). The 2-year cumulative NRM was 24.5% and 7.1%, respectively (P=0.120). The 2-year LFS was 58.5% and 50.0%, respectively (P=0.466). The 2-year OS rate was 69.8% and 64.3%, respectively (P=0.697). In the multivariate analysis, the conditioning regimen containing TBI was a protective factor for relapse after transplantation (HR=0.304, 95% CI 0.135-0.688, P=0.004), whereas the effect on NRM was not significant (HR=1.393, 95% CI 0.355-5.462, P=0.634). Infection was an independent risk factor for OS after allo-HSCT in adult patients with ALL. Conclusion: allo-HSCT based on TBI conditioning regimen had lower relapse rate and lower incidence of hemorrhagic cystitis for adult ALL, compared with chemotherapy regimen. While the incidence o grade Ⅲ/Ⅳ aGVHD was hgher in TBI conditioning regimen than that in chemotherapy regimen.

Myb overexpression synergizes with the loss of Pten and is a dependency factor and therapeutic target in T-cell lymphoblastic leukemia.

T-lineage acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy that accounts for 10%-15% of pediatric and 25% of adult ALL cases. Although the prognosis of T-ALL has improved over time, the outcome of T-ALL patients with primary resistant or relapsed leukemia remains poor. Therefore, further progress in the treatment of T-ALL requires a better understanding of its biology and the development of more effective precision oncologic therapies. The proto-oncogene MYB is highly expressed in diverse hematologic malignancies, including T-ALLs with genomic aberrations that further potentiate its expression and activity. Previous studies have associated MYB with a malignant role in the pathogenesis of several cancers. However, its role in the induction and maintenance of T-ALL remains relatively poorly understood. In this study, we found that an increased copy number of MYB is associated with higher MYB expression levels, and might be associated with inferior event-free survival of pediatric T-ALL patients. Using our previously described conditional Myb overexpression mice, we generated two distinct MYB-driven T-ALL mouse models. We demonstrated that the overexpression of Myb synergizes with Pten deletion but not with the overexpression of Lmo2to accelerate the development of T-cell lymphoblastic leukemias. We also showed that MYB is a dependency factor in T-ALL since RNA interference of Myb blocked cell cycle progression and induced apoptosis in both human and murine T-ALL cell lines. Finally, we provide preclinical evidence that targeting the transcriptional activity of MYB can be a useful therapeutic strategy for the treatment of T-ALL.

Bridging Therapy in Adults with Relapsed/Refractory B-Cell Acute Lymphoblastic Leukemia Receiving Standard of Care Brexucabtagene Autoleucel: Results from the Real-World Outcomes Collaborative of CAR T in Adult ALL (ROCCA)

Bridging Therapy in Adults with Relapsed/Refractory B-Cell Acute Lymphoblastic Leukemia Receiving Standard of Care Brexucabtagene Autoleucel: Results from the Real-World Outcomes Collaborative of CAR T in Adult ALL (ROCCA)

The BET inhibitor GNE-987 effectively induces anti-cancer effects in T-cell acute lymphoblastic leukemia by targeting enhancer regulated genes.

T-cell acute lymphoblastic leukemia (T-ALL) is a highly aggressive hematologic malignancy originating from T progenitor cells. It accounts for 15% of childhood and 25% of adult ALL cases. GNE-987 is a novel chimeric molecule developed using proteolysis-targeting chimeras (PROTAC) technology for targeted therapy. It consists of a potent inhibitor of the bromodomain and extraterminal (BET) protein, as well as the E3 ubiquitin ligase Von Hippel-Lindau (VHL), which enables the effective induction of proteasomal degradation of BRD4. Although GNE-987 has shown persistent inhibition of cell proliferation and apoptosis, its specific antitumor activity in T-ALL remains unclear. In this study, we aimed to investigate the molecular mechanisms underlying the antitumor effect of GNE-987 in T-ALL. To achieve this, we employed technologies including RNA sequencing (RNA-seq), chromatin immunoprecipitation sequencing (ChIP-seq) and CUT&Tag. The degradation of BET proteins, specifically BRD4, by GNE-987 has a profound impact on T-ALL cell. In in vivo experiments, sh-BRD4 lentivirus reduced T-ALL cell proliferation and invasion, extending the survival time of mice. The RNA-seq and CUT&Tag analyses provided further insights into the mechanism of action of GNE-987 in T-ALL. These analyses revealed that GNE-987 possesses the ability to suppress the expression of various genes associated with super-enhancers (SEs), including lymphoblastic leukemia 1 (LCK). By targeting these SE-associated genes, GNE-987 effectively inhibits the progression of T-ALL. Importantly, SE-related oncogenes like LCK were identified as critical targets of GNE-987. Based on these findings, GNE-987 holds promise as a potential novel candidate drug for the treatment of T-ALL.

The landscape of alterations affecting epigenetic regulators in T-cell acute lymphoblastic leukemia: Roles in leukemogenesis and therapeutic opportunities.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy accounting for 10%-15% of pediatric and 20%-25% of adult ALL cases. Epigenetic irregularities in T-ALL include alterations in both DNA methylation and the post-translational modifications on histones which together play a critical role in the initiation and development of T-ALL. Characterizing the oncogenic mutations that result in these epigenetic changes combined with the reversibility of epigenetic modifications represents an opportunity for the development of epigenetic therapies. Oncogenic mutations and deregulated expression of DNA methyltransferases (DNMTs), Ten-Eleven Translocation dioxygenases (TETs), Histone acetyltransferases (HATs) and members of Polycomb Repressor Complex 2 (PRC2) have all been identified in T-ALL. This review focuses on the current understanding of how these mutations lead to epigenetic changes in T-ALL, their association with disease pathogenesis and the current efforts to exploit these clinically through the development of epigenetic therapies in T-ALL treatment.

Long-Term Results of the Dasatinib-Blinatumomab Protocol for Adult Philadelphia-Positive ALL.

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned coprimary or secondary analyses are not yet available. Clinical trial updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported.We report the long-term results of the frontline trial with dasatinib and blinatumomab in induction/consolidation (GIMEMA LAL2116, D-ALBA) for adult Philadelphia-positive ALL (Ph+ ALL), which enrolled 63 patients of all ages. At a median follow-up of 53 months, disease-free survival, overall survival, and event-free survival are 75.8%, 80.7%, and 74.6%, respectively. No events have occurred among early molecular responders. A significantly worse outcome was recorded for IKZF1plus patients. Twenty-nine patients-93.1% being in molecular response (ie, complete molecular response or positive nonquantifiable) after dasatinib/blinatumomab-never received chemotherapy/transplant and continued with a tyrosine kinase inhibitor only; 28 patients remain in long-term complete hematologic response (CHR). An allogeneic transplant was carried out in first CHR mainly in patients with persistent minimal residual disease; 83.3% of patients are in continuous CHR. The transplant-related mortality was 12.5% for patients transplanted in first CHR and 13.7% overall. Nine relapses and six deaths have occurred. ABL1 mutations were found in seven cases. The final analysis of the D-ALBA study shows that a chemotherapy-free induction/consolidation regimen on the basis of a targeted strategy (dasatinib) and immunotherapy (blinatumomab) is effective in inducing durable long-term hematologic and molecular responses in adult Ph+ ALL, paving the way for a new era in the management of these patients.

Efficacy and Safety of Chimeric Antigen Receptor T Cells Therapy Strategy That Dual Targeting CD19 and CD22 to Treat Relapsed/Refractory Acute Lymphoblastic Leukemia in Adults

Background: Chimeric antigen receptor T-cells (CAR-T) targeting CD19 therapy is reported to induce 83-93% response in relapsed/refractory acute lymphoblastic leukemia (r/r-ALL) in adults. However, 30-60% of patients relapse after CAR-T treatment, of which 14-25% CD19-negative relapse. In spite of CAR-T cells persistence, CD19 absent causing tumors that evade CAR-T cells mediated recognition and clearance. For CD19-negative relapse, it is necessary to explore more effective targets. There are some research proved that compared with single-targeted CARs, CD19/CD22 dual-targeted CARs induce more IFN-γ and IL-2 in vitro and eradicate patient-derived xenografts (PDX) produced with CD19-negative relapse of CD19-directing CAR-T treatment. Therefore, our study explored the efficacy and safety of clinical studies on mixture of CD19 CAR-T and CD22 CAR-T cells in the treatment of r/r ALL in adults. Patients and Methods: CD3 + cells were selected from the apheresis PBMC and activated before lentiviral CAR T cells infection. The cells were transduced with a caspase9-inducible, safety-engineered lentivector CAR containing anti-CD19 or -CD22 scFv fused with multiple intracellular signaling domains: CD28/CD27/CD3z-iCasp9. The quality of apheresis cells, gene transfer and T cell proliferation efficiencies, and effective CAR-T infusion dose were quantitatively scored and documented. All enrolled patients expressed CD19 and CD22 positively. All patients received fludarabine (FLU) and cyclophosphamide (CTX) conditioning chemotherapy (FLU 30mg/m 2, d1-3; CTX 500mg/m 2, d1-3) before CAR-T infusions. Results: The study included 16 r/r ALL patients, median age is 26 years, and median of previous theraples is 6, all of patient details can be found in the table (Fig.1). In the first month after CD19/CD22 CAR-T cells infusion, total of 93.8% (15/16) of patients achieved complete response (CR), 1 case (6.25%) was evaluated as non-remission (NR). In the evaluation of safety, 68.75% (11/16) of patients did not experience cytokine release syndrome (CRS), 25% of patients (4/16) occurred grade 1 CRS and 6.25% (1/16) of patients occurred grade 2 CRS, no case of severe CRS defined as≥grade 3 and no any immune effector cell-associated neurotoxicity syndrome (ICANS) occurred. After a median follow-up of 9.9 months, 4 patients maintained CR, while other 11 patients were relapsed, with a duration of response (DOR) of 25% (4/16), median LFS was 6.1 months and median OS were not reached (Fig.2). Conclusions: In summary, CD19/CD22 exhibited a manageable safety but limited antileukemia activity. Our study showed that mixture of CD19 CAR-T and CD22 CAR-T cells infusion is fail to expected to compensate for the treatment of relapsed in r/r ALL and prolonging the leukemia-free survival of patients. The low-efficacy of dual-targeted CAR-T cells may be caused by the CAR19 and CAR22 cells interfere with each other and weaken the CAR-T amplification in vivo. From our data, it seems that the timing of treatment and tumor burden status may play a more important role rather than new targets CAR-T cells infusion for r/r ALL in adults.

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).

Olverembatinib (HQP1351) Enhances Antitumor Effects of Chemotherapy in Philadelphia Chromosome-Positive (Ph⁺) Acute Lymphoblastic Leukemia (ALL)

Introduction Ph⁺ ALL is the most common subtype of B-cell- (B-) ALL in adults. BCR-ABL1 tyrosine kinase inhibitors (TKIs) are effective against chronic myeloid leukemia (CML) cells. However, treatment with TKIs fails to maintain disease remission in Ph⁺ B-ALL. Although the complete hematologic remission rate is very high, cytogenetic and molecular remission rates are low, so that TKIs are still considered as complements to chemotherapy and bridges to allogeneic stem cell transplantation. Olverembatinib is a novel, third-generation TKI with more potent activity against BCR-ABL1 and overcomes various resistant mutations, including the “gatekeeper” T315I, which confers resistance against all first- and second-generation TKIs. The aim of this study was to assess whether olverembatinib combined with chemotherapeutic agents vincristine or doxorubicin can promote anti-cancer effects in Ph⁺ ALL cells and explore the potential mechanism of action of the combinations. Methods Cell-based antiproliferation assays to evaluate the activities of olverembatinib alone or in combination with vincristine or doxorubicin were conducted using the CellTiter-Glo® Luminescent Cell Viability assay kits. Drug induced cell apoptosis was measured by flow cytometry. Potential mechanism of action analysis was conducted using western blot to evaluate the protein expression involved in BCR-ABL1 and apoptosis signaling pathways. Results Cell-based antiproliferation studies showed synergistic activity of olverembatinib in combination with vincristine or doxorubicin in Ph⁺ ALL SUP-B15 cells compared with single agents after 72 hours of treatment. The percentage of apoptotic cells induced by 24-hour treatment of olverembatinib (100 nM), vincristine (300 nM), and the combination were 27.7%, 27.2%, and 40.7%, respectively, while the percentage of apoptotic cells induced by 24-hour treatment of doxorubicin (100 nM) alone and in combination with olverembatinib (100 nM) were 35.3 and 74.1, respectively (apoptotic cells % in DMSO control group is 6.74). The combination significantly increased cell apoptosis compared with single agents ( p &amp;lt; 0.0001). Mechanistically, the combination of olverembatinib with vincristine showed synergistic effects in inhibiting protein expression and phosphorylation of BCR-ABL1, and downstream signal transducer and activator of transcription 5 (STAT5). The synergistic effects were also seen in decreased phosphorylation of AKT and ERK1/2. In addition, the combination significantly decreased protein levels of antiapoptotic MCL-1, BCL-2, and BCL-xL and increased protein levels of proapoptotic BAX and p53 upregulated modulator of apoptosis (PUMA) in a dose-dependent manner, which led to marked augmentation of cleavage of caspase-3 and poly (ADP-ribose) polymerase 1 (PARP). A similar pattern was also observed when olverembatinib was combined with doxorubicin. Conclusions Olverembatinib, a novel, third-generation TKI, in combination with chemotherapeutic agents vincristine or doxorubicin decreased MCL-1, BCL-2, and BCL-xL and increased BAX and PUMA in a dose-dependent manner, the combinations have demonstrated synergistic antitumor effects by enhancing apoptosis and antiproliferation in Ph⁺ ALL cells, which may provide an alternative approach for treatment of patients with Ph⁺ ALL.

Jmjd3 Drives Immature T-Cell Lymphoblastic Leukemia

Introduction: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy that accounts for 15% of pediatric and 25% of adult ALL cases. Despite an improved survival rate due to intensified chemotherapy and bone marrow transplantation, still 20% of T-ALL patients are primary refractory to the standard treatment or relapse, and have a poor clinical outcome. T-ALL patients can be classified into molecular genetic subgroups based on the expression or genomic alterations of specific oncogenic transcription factors. One such subgroup the early T-cell precursor ALL (ETP-ALL) is a heterogeneous subgroup of immature leukemias that often display high expression of LYL1 and LMO2. It's important to improve our knowledge of T-ALL biology and developing novel targeted anti-TALL therapies. We previously showed that the H3K27 demethylase Jmjd3 plays an important role in T-ALL and is a good candidate for the treatment of T-cell leukemia. Aim: Identify the role of Jmjd3 in the initiation of immature T-ALL. Methods: We generated a conditional Rosa26- Jmjd3 overexpression mouse model (Figure 1A), and crossed it with a hematopoietic-specific Vav-icre mice, to obtain Jmjd3 Vav mice. To investigate the role of Jmjd3 in tumor initiation, we monitored survival of a Jmjd3 Vav cohort. Arising tumors were characterized by immunohistochemistry, flow cytometry, and were transplanted into secondary immunodeficient recipients. To characterize the pre-leukemic compartment, we performed single cell feature barcoding and RNA sequencing. Also bulk RNAseq and proteomics were performed on primary and transplanted Jmjd3-driven tumors. To investigate the role of Jmjd3 in immature T-cell biology in human and mice, we analyzed thymi from 8-weeks-old control and Jmjd3 Vavmice, and performed OP9-DL4 stromal co-cultures with transduced CD34+ human cord blood cells that overexpressed JMJD3, respectively. We used ETP-ALL patient-derived xenografts (PDX) models to validate markers identified in single cell analysis. Results: We found that hematopoietic Jmjd3 overexpression resulted in hematological malignancies with 38% penetrance (n= 8/21) and a median survival of 63 weeks (Figure 1B). Immunohistochemistry confirmed overexpression of Jmjd3, decreased levels of H3K27me3, and increased levels of H3K27ac in Jmjd3 Vav tumors. Immunophenotypic analysis revealed immature Thy1.1 +CD4 -CD8 - cells in all obtained Jmjd3 Vav tumors (Figure 1C), indicative of an immature the early T-cell precursor (ETP)-like ALL. These immature T-ALLs gave rise to primary (7 weeks) and secondary (3 weeks) transplants upon their transplantation in immunocompromised mice. Bulk RNAseq confirmed that the transcriptional feature of the primary tumor were maintained upon transplantation. The role of Jmjd3 in stimulating immature thymocyte development was confirmed in thymi from 8-weeks-old Jmjd3 Vavmice and could be validated as well in human T-cell development using stromal OP9-DL4 co-cultures. Finally, we used single cell sequencing technologies to characterize both transcriptional features and cell-surface markers within the tumor fraction (Figure 1D). We confirmed that the tumor cluster was ETP-like with absence of CD1, CD5 and mature T-cell markers, and expression of CD44, Lyl1, and myeloid markers. Furthermore, we could identify cell-surface in immature Jmjd3-driven tumors that were also present in human T-cell stromal culture and ETP-ALL PDXs. Conclusion: We identified Jmjd3 as a driver of immature thymocyte populations that give rise to ETP-ALL.

Diagnostic Utility of Comprehensive RNA-Seq Analysis in Adult B-ALL

Introduction Adult B-cell acute lymphoblastic leukemia (B-ALL) comprises 75% of adult ALL cases. The classification and risk stratification of patients have conventionally relied on cytogenetics analysis. However, cytogenetics has limitations due to its reliance on metaphase cells, which can mitigate classification accuracy. In recent years, transcriptome sequencing (RNA-seq) of large pediatrics ALL cohorts has expanded the number of recurrent gene fusions recognized as disease defining events and informed risk stratification [Brady et al. 2022, Mullighan et al. 2017]. This study aims to evaluate the diagnostic utility of RNA-seq for clinical subtype classification in adult B-ALL in a large representative cohort. Methods The study cohort comprised 338 adult patients (25-65 years) with newly diagnosed ALL (UKALL14, ISRCTN66541317, NCT01085617). Paired-end RNA-seq data at 60 Million reads per sample were generated. Analysis focused on the detection of disease-defining gene fusions and gene expression signatures. Three RNA fusion callers (FusionCatcher, FuSeq, and STAR-Fusion) and three expression-based classifiers (ALLSorts, ALLspice, and ALLCatchR) were used for consensus fusion calling and expression-based classification, respectively [Nicorici et al. 2014, Vu et al. 2018, Haas et al. 2019, Schmidt et al., Mäkinen et al., Bader et al. 2022]. Integration of classification results from expression-based classifiers and the consensus gene fusions provides 4 levels of RNA-derived subtype information, denoted as “RNA evidence” in this study. These RNA-seq based classification results were compared with clinical cytogenetics findings. Results Out of 338 samples in this cohort, cytogenetics unequivocally classified 73% (246/338) of the samples. By integrating RNA expression-based classification with fusion subtypes, we confidently classified 87% (295/338) of the cohort with at least 2 levels of RNA evidence. In total, consensus gene fusions were identified in 74% (249/338), while expression-based classifiers assigned a confident subtype to 82% (319/338). Subsequently, we compared cytogenetics with RNA-seq derived classification and found a high concordance of 98% (240/246) between the two approaches. Only 6 (&amp;lt;2%) samples with cytogenetics subtypes were not classified by RNA-seq, attributed to specific subtype characteristics (near-haploid) or lower RNA purity/quality. 27% (92/338) of subjects were not classified by cytogenetics (31 missing/failed and 61 indeterminate cytogenetics). Of the 31 samples with failed/missing cytogenetics, RNA-seq confidently assigned a class in 74% (23/31). For the remaining 61 cases with indeterminate cytogenetic (complex/other), RNA-seq resolved classification in 74% (45/61). Taken together, this improved the classification for 20% (68/338) of the entire cohort that was previously unclassified through cytogenetics. Notably, RNA-seq-exclusive findings were enriched in subtypes like DUX4, PAX5alt, and ZNF384, which are challenging for cytogenetics to discern. Importantly, integration of RNA-seq with cytogenetics led to a confident subtype classification for 93% of the cohort (314/338), a significant improvement over the initial 73% based on cytogenetics alone. Conclusions In conclusion, our study highlights the relevance of integrated RNA-seq analysis for reliable and comprehensive subtype classification in adult B-ALL. The high concordance between cytogenetics and RNA-derived subtypes underscores their complementary roles, providing valuable insights into disease mechanisms and guiding personalized treatment strategies.

CNS Irradiation in Adult De Novo B-Precursor ALL / Lbl: Results of the Randomized GMALL T rial 08/2013 Indicate Potential Antileukemic Efficacy

In the past decade outcome of adult ALL was improved significantly using pediatric-based chemotherapy. The GMALL Trial 08/2013 (NCT02881086) for patients (pts) aged 18-55 years (yrs) with newly diagnosed ALL/LBL is a pediatric based regimen with MRD based treatment stratification and risk adapted stem cell transplantation (SCT) (Gökbuget et al, ASH 2021). CNS prophylaxis consisted of three doses i.th. MTX during induction II, 11 doses of triple i.th. for standard risk (SR) in consolidation (C)/maintenance and 1 dose before SCT in high risk (HR); 7 vs 1 cycle of HDMTX (1.5 g/m 2 as 24h infusion) were scheduled in SR and HR resp. Prophylactic fractionated CNS irradiation (RAD) with 24 Gy during induction II (IP2) was part of all previous GMALL trials. The trial incorporated a randomization for Ph/BCR::ABL1 negative (Ph-) B-precursor ALL/LBL without initial CNS involvement and treated until C1 in order to compare standard IP2 with i.th. MTX with vs without CNS RAD. The key endpoint was the time from start of induction to start of C1 in CR pts, reasoning that less toxicities and treatment delays would occur without CNS RAD. Further endpoints were remission duration (RD), event-free survival (EFS) and cognitive disturbances (DemTect Test). The statistical assumption was that it is possible to detect a shortening of time to C1 from 74 to 69 days with a power of more than 90% with 215 pts per arm with a two-sided α-niveau of 5%. Between 8/2016-8/2022 1023 pts from 78 centers were included. 512 pts had Ph- B-precursor ALL/LBL (497/15). 31 pts had initial CNS involvement (N=20) or were not randomized due to other reasons (N=11). 241 pts were randomized in arm A (RAD) and 241 in arm B (NoRAD). The median age was 33.5 (18-55) yrs; c/pre B-ALL versus pro B-ALL was present in 83% vs 17%. 191 were HR vs 291 SR. No significant differences were observed between both arms. 477 pts were randomized (ITT population) and 422 were eligible (analysis population) since they achieved CR and were treated until C1. Reasons for non-eligibility were protocol violation (N=5), early death (ED) (N=15), withdrawal (N=12), no CR before C1 (N=18) and late withdrawals in CR before C1 (N=10). 207 pts in the RAD-arm (86%) vs 215 pts in the NoRAD-arm (89%) were eligible. In the analysis population treatment per randomization was 97% in NoRAD and 95% in RAD arm. CR rates before C1 (95% RAD vs 96% NoRAD), ED rates (5% vs 4%), rates of grade III/IV leukopenia in IP2 (14% vs 16%) and infections (29% vs 29%) were similar. The median time to C1 was similar too (75 vs 76 days; p=0.34). Relapse rates (RR) were lower in RAD (13%) vs NoRAD (18%) (p=0.14). Whereas CNS relapses were infrequent in both arms (N=1 vs N=3 i.e. 1% vs 1%; p=0.62), the rate of BM±other relapses (11% vs 15%; p=0.15) was lower in the RAD arm. The cumulative incidence of CNS relapse at 3 yrs was 1% vs 1% and of all relapses 14% vs 20% (p=0.13). The EFS at 3 yrs was higher for the RAD (79% [95%-CI: 72% - 85%]) vs NoRAD (72% [95%-CI: 65% - 78%])(p=0.10) in the analysis population and 74% vs 67% (p=0.13) in the ITT population. In the ITT population the impact was statistically significant (p=0.05) in SR pts whereas no difference was detected in HR (i.e. pts with indication for SCT) (figure 1); differences did not reach significance in the analysis population. We further analyzed the impact of randomization arm on cognitive function (analysis population). After C1 results were available in 264 pts. In the RAD vs NoRAD arm 2% vs 2% resp. had low scores (0-8) and 10% vs 8% intermediate scores (9-12) indicating a relevant or limited cognitive disturbance. At the end of therapy (week 130), results are available in 53 pts treated in SR arm. 0% vs 4% had low and 4% vs 4% intermediate scores. Thus, relevant cognitive limitations were present in both arms. The primary endpoint (time to C1) was not met. Pts with RAD did have a trend towards better EFS and RD. The CNS RR was overall low but CNS RAD might have an impact on the systemic relapse rate in SR pts. We recorded no significant negative impact of CNS RAD on DemTect results, but this requires further observation. Results must be interpreted in the context of adult pts with potentially lower risks for cognitive disturbances compared to children and - on the other hand - a relatively low dose of systemic HDMTX. Overall, no clear negative effects of CNS RAD were detected whereas a trend towards positive impact on EFS was observed. GMALL 08/2013 is an independent academic trial funded by the Deutsche Krebshilfe.