Adult T-cell Acute Lymphoblastic Leukemia Research Articles

Long-term exposure and response to azacitidine for post-hematopoietic stem cell transplantation relapse of early T-cell precursor acute lymphoblastic leukemia: a case report and review of the literature

Adult T-cell acute lymphoblastic leukemia has a poor outcome after relapse. Because the subtype of early T-cell precursor displays characteristics close of those of acute myeloid leukemia, such as epigenetic dysregulation, hypomethylating agents might prove of interest. We describe the case of a patient relapsing 3 months only after allogeneic stem cell transplantation who achieved complete remission on azacitidine, and is still on therapy 9 years later. We discuss the biological background of this very long-term response, underlining the immunological effects of hypomethylating agents, and the perspectives opened by combination of hypomethylating agents with other drugs such as venetoclax.

Contribution of copy number to improve risk stratification of adult T-cell acute lymphoblastic leukemia patients enrolled in measurable residual disease-oriented trials.

Not available.

NGS-based stratification refines the risk stratification in T-ALL and identifies a very-high-risk subgroup of patients

AbstractWe previously reported a better outcome in adult and pediatric T-cell acute lymphoblastic leukemia (T-ALL) harboring NOTCH1 and/or FBXW7 mutations without alterations of K-N-RAS and PTEN genes. Availability of high-throughput next-generation sequencing (NGS) strategies led us to refine the outcome prediction in T-ALL. Targeted whole-exome sequencing of 72 T-ALL–related oncogenes was performed in 198 adults with T-ALLs in first remission from the GRAALL-2003/2005 protocols and 242 pediatric patients with T-ALLs from the FRALLE2000T. This approach enabled the identification of, to our knowledge, the first NGS-based classifier in T-ALL, categorizing low-risk patients as those with N/F, PHF6, or EP300 mutations, excluding N-K-RAS, PI3K pathway (PTEN, PIK3CA, and PIK3R1), TP53, DNMT3A, IDH1/2, and IKZF1 alterations, with a 5-year cumulative incidence of relapse (CIR) estimated at 21%. Conversely, the remaining patients were classified as high risk, exhibiting a 5-year CIR estimated at 47%. We externally validated this stratification in the pediatric cohort. NGS-based classifier was highly prognostic independently of minimal residual disease (MRD) and white blood cell (WBC) counts, in both adult and pediatric cohorts. Integration of the NGS-based classifier into a comprehensive risk-stratification model, including WBC count at diagnosis and MRD at the end of induction, enabled the identification of an adverse-risk subgroup (25%) with a 5-year CIR estimated at 51%, and a favorable-risk group (32%) with a 5-year CIR estimated at 12%. NGS-based stratification combined with WBC and MRD sharpens the prognostic classification in T-ALL and identifies a new subgroup of patients who may benefit from innovative therapeutic approaches. The GRAALL-2003/2005 studies were registered at www.ClinicalTrials.gov as #NCT00222027 and #NCT00327678.

Prognostic factors impacting post-transplant outcomes in adult T-cell acute lymphoblastic leukemia: a registry-based study by the EBMT acute leukemia working party.

T-cell acute lymphoblastic leukemia (T-ALL) predominantly affects individuals in late childhood and young adulthood. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative modality particularly in the setting of poor risk genetics and/or persistent minimal residual disease. Limited studies have directly explored the impact of patient- and transplant-related factors on post-transplant outcomes in T-ALL. Using a large dataset from the European Society for Blood and Marrow Transplantation registry, we identified 1907 adult T-ALL patients (70% male) who underwent their first allo-HSCT in first complete remission (CR1) from matched sibling donors (MSD; 45%), unrelated donors (UD; 43%) or haploidentical donors (12%) between 2010 and 2021. The median age at transplant was 33.4 years (18.1-75). The median follow up was 2.9 years. Most patients underwent total body irradiation (TBI)-based myeloablative conditioning (69%). The 2-year overall survival (OS) was 69.4%, and leukemia -free survival (LFS) was 62.1%. In multivariate analysis, advanced age at transplant negatively affected LFS (for each 10-year increment, HR = 1.11, p = 0.004), GVHD-free, relapse-free survival (GRFS) (HR = 1.06, p = 0.04), OS (HR = 1.12, p = 0.002), and non-relapse mortality (NRM) (HR = 1.23, p < 0.001). More recent years of allo-HSCT were associated with improved GFRS (For each 3-year increment, HR = 0.89, p < 0.001), OS (HR = 0.9, p = 0.02), and decreased NRM (HR = 0.82, p = 0.008). TBI improved LFS. (HR = 0.79, p = 0.02), GRFS (HR = 0.83, p = 0.04), and relapse incidence (RI) (HR = 0.65, p < 0.001). Female-to-male transplant negatively affected GRFS (HR = 1.21, p = 0.02) and OS (HR = 1.23, p = 0.048). In vivo T-cell depletion significantly improved GFRS (HR = 0.74, p < 0.001). This large study identified prognostic factors, such as age at transplant conditioning regimen, in influencing post-transplant in adult T-ALL patients undergoing allo-HSCT. Importantly, a significant improvement over time was noted. These findings hold great promise for new adapted treatment strategies and can serve as a benchmark for future studies in that setting.

The glutamate/aspartate transporter EAAT1 is crucial for T-cell acute lymphoblastic leukemia proliferation and survival.

T-cell acute lymphoblastic leukemia (T-ALL) is a cancer of the immune system. Approximately 20% of paediatric and 50% of adult T-ALL patients have refractory disease or relapse and die from the disease. To improve patient outcome new therapeutics are needed. With the aim to identify new therapeutic targets, we combined the analysis of T-ALL gene expression and metabolism to identify the metabolic adaptations that T-ALL cells exhibit. We found that glutamine uptake is essential for T-ALL proliferation. Isotope tracing experiments showed that glutamine fuels aspartate synthesis through the TCA cycle and that glutamine and glutamine-derived aspartate together supply three nitrogen atoms in purines and all but one atom in pyrimidine rings. We show that the glutamate-aspartate transporter EAAT1 (SLC1A3), which is normally expressed in the central nervous system, is crucial for glutamine conversion to aspartate and nucleotides and that T-ALL cell proliferation depends on EAAT1 function. Through this work, we identify EAAT1 as a novel therapeutic target for T-ALL treatment.

Significance of immunophenotypic, cytogenetic, and molecular markers in adult patients with T-cell lymphoblastic leukemia

Background. Current chemotherapy protocols for T-cell acute lymphoblastic leukemia (T-ALL) allow achieving a 5-year overall survival of 60–90 %, but relapsed and refractory forms remain incurable situations.Aim. To determine the significance of immunophenotypic, cytogenetic and molecular markers in adult T-ALL patients receiving therapy according to the ALL-2016 protocol.Materials and methods. From December 2016 to June 2022, 113 patients with primary T-ALL were included in the study. Cytogenetic study was performed in 104 (92 %) patients; anomalies in the IKZF1 and NOTCH1 genes were investigated in 43 (38 %) patients.Results. The worst prognosis was in patients with ETP and near-ETP variants of T-ALL (3-year disease-free survival was 54 % in ETP group, 33 % in near-ETP group vs TI/II – 79 %, TIII – 89 %, TIV – 75 %). In early T-ALL variants, abnormal karyotype was most common (ETP – 80.7 %, near-ETP – 60 %). Aberrations in NOTCH1 gene were found in 53 % of cases (in 23 out of 43 patients), and no mutations were found in IKZF1 gene in our study. In the group with no NOTCH1 abnormalities, the overall survival was significantly worse than in the group with abnormalities (NOTCH1– – 52 % vs NOTCH1+ –81 %; p = 0.05).

Genomic imbalance analysis provides new insight into prognostic factors in adult and pediatric T-ALL

AbstractGiven the poor outcome of refractory and relapsing T-cell acute lymphoblastic leukemia (T-ALL), identifying prognostic markers is still challenging. Using single nucleotide polymorphism (SNP) array analysis, we provide a comprehensive analysis of genomic imbalances in a cohort of 317 newly diagnosed patients with T-ALL including 135 children and 182 adults with respect to clinical and biological features and outcomes. SNP array results identified at least 1 somatic genomic imbalance in virtually all patients with T-ALL (∼96%). Del(9)(p21) (∼70%) and UPD(9)p21)/CDKN2A/B (∼28%) were the most frequent genomic imbalances. Unexpectedly del(13)(q14)/RB1/DLEU1 (∼14%) was the second most frequent copy number variant followed by del(6)(q15)/CASP8AP2 (∼11%), del(1)(p33)/SIL-TAL1 (∼11%), del(12)(p13)ETV6/CDKN1B (∼9%), del(18)(p11)/PTPN2 (∼9%), del(1)(p36)/RPL22 (∼9%), and del(17)(q11)/NF1/SUZ12 (∼8%). SNP array also revealed distinct profiles of genomic imbalances according to age, immunophenotype, and oncogenetic subgroups. In particular, adult patients with T-ALL demonstrated a significantly higher incidence of del(1)(p36)/RPL22, and del(13)(q14)/RB1/DLEU1, and lower incidence of del(9)(p21) and UPD(9p21)/CDKN2A/B. We determined a threshold of 15 genomic imbalances to stratify patients into high- and low-risk groups of relapse. Survival analysis also revealed the poor outcome, despite the low number of affected cases, conferred by the presence of chromothripsis (n = 6, ∼2%), del(16)(p13)/CREBBP (n = 15, ∼5%) as well as the newly-identified recurrent gain at 6q27 involving MLLT4 (n = 10, ∼3%). Genomic complexity, del(16)(p13)/CREBBP and gain at 6q27 involving MLLT4, maintained their significance in multivariate analysis for survival outcome. Our study thus demonstrated that whole genome analysis of imbalances provides new insights to refine risk stratification in T-ALL. This trial was registered at www.ClinicalTrials.gov as #NCT00222027 and #NCT00327678, and as #FRALLE 2000T trial.

NGS-Based Stratification Refines the Risk Stratification in T-ALL and Identifies a Very High-Risk Subgroup of Patients

Purpose We previously reported a significantly better outcome in adult and pediatric T-cell acute lymphoblastic leukemia (T-ALL) harboring NOTCH1 and/or FBXW7 ( N/F) mutations without alterations of K-N-RAS and PTEN(R/P) genes . High-throughput next-generation sequencing strategies (NGS) allowed us to refine the prediction of outcome in T-ALL. Patients and Methods 198 adult T-ALLs in first remission (CR1) from the GRAALL-2003/2005 protocol were included in the study as the construction cohort, and 242 pediatric T-ALLs from FRALLE2000 were used as a validation cohort. Targeted whole-exome sequencing of 63 T-ALL-related oncogenes was performed. Primary outcome was cumulative incidence of relapse (CIR). To account for the large number of candidates genes, selection was performed using a LASSO penalization in a Fine and Gray model predicting CIR (Fu Z. et al. Lifetime Data Anal., 2017). To construct the final risk-stratification score, we used non-parametric clustering of CIR curves through k-medians algorithm (Villanueva N. et al. Stat. Med., 2019). Results We confirm the prognostic classifier NFRP in the NGS era and evaluate the impact of 39 new gene alterations in the adult cohort. Alterations affecting TP53, DNMT3A, IDH1/2, IKZF1, PI3K pathway ( PTEN, PIK3CA and PIK3R1), EP300, and PHF6 were independent prognostic factors in adult T-ALL. This led us to propose the first NGS-based classifier in T-ALL defining low risk patient (LR) as those with N/F, PHF6 or EP300 mutations without N-K-RAS, PI3K pathway , TP53, DNMT3A, IDH1/2 and IKZF1 alterations (234 of 440 patients, 53%). In the adult cohort, the NGS-based classifier separates a high-risk group (HR) (n=90/198, 45%) with a 5-year CIR of 47% (95%CI:36%-57%) and a low-risk group (LR) (n=108/198, 55%) with a 5-year CIR of 21% (95%CI:14%-29%) (p&amp;lt;0.0001). Our NGS-classifier was validated in the pediatric cohort, with a 5-year CIR of 35% (95%CI:26%-44%) in HR group (n=116/242, 48%) and 5-year CIR of 17% (95%CI:11%-24%) in the LR group (n=126/242, 52%) (p=0.001) ( Figure A). Since the NGS-based classifier is highly prognostic independently of minimal residual disease (MRD) at end of induction (cutoff 10 -4) and white blood cells count (WBC) (cutoff 100 x 10 9/L), we then developed and externally validated a global risk stratification model incorporating MRD1, WBC at diagnosis and the NGS-classifier. This model identifies 3 subgroups at CR1: a large favorable Risk (CR1-FAV) group (231/332, 70%) with CIR at 5 years estimated at 19% (95%CI:14%-24%) ( Figure B), a subgroup of Adverse risk (CR1-ADV) patients (30/332, 9%) with a 5-year CIR of 68% (95%CI:46%-82%) and an Intermediate risk (CR1-INT) group (71/332, 21%) with a 5-year CIR of 37% (95%CI:26%-48%). Conclusion T-ALL NGS-based stratification combined with WBC and MRD evaluation sharpens the prognostic classification in T-ALL and identifies a new subgroup of adverse risk patients who should benefit from innovative therapeutic approaches.

Prognostic Factors Impacting Post-Transplant Outcomes in Adult T-Cell Acute Lymphoblastic Leukemia: A Registry-Based Study By the EBMT Acute Leukemia Working Party

Background: T-cell acute lymphoblastic leukemia (T-ALL) predominantly affects individuals in late childhood and young adulthood and historically had high mortality rates. The outcome of Pediatric inspired chemo protocols in the T-ALL have improved in the past years due to the advancement in treatment and supportive care but the data is less mature, although adult patients still face challenges. Limited studies have directly explored the impact of patient- and transplant-related factors on post-transplant outcomes in adult T-ALL patients after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Methods: A retrospective registry-based analysis was conducted across multiple centers using data from the European Society for Blood and Marrow Transplantation (EBMT) registry. Inclusion criteria were adult T-ALL patients (&amp;gt;18 years old) who underwent their first allo-HSCT in first complete remission from matched sibling donors (MSD), unrelated donors (UD) (10/10 or 9/10) or haploidentical donors between 2010 and 2021. Cord bloods were excluded. Multivariable Cox regression analysis was performed to examine the associations between patient/transplant characteristics and outcomes. Results: A total of 1907 patients were included in the study. The median age at transplant was 33.4 years (18.1-75). The median follow up was 2.9 years (2.6-3.1), and the median year of HSCT was 2016 (2010-2021). Patients were 70 % male with 67% being CMV-positive, at the time of transplant. Donors were predominantly male (64%), and (23%) of transplants involved female donors to male patients. The median period between diagnosis and HSCT was 5.9 months (0 -23.8). Most transplants were from matched sibling donors (MSD) (45%) followed by UD (43%) and 12% were from Haploidentical transplants. Most patients underwent myeloablative conditioning with total body irradiation (TBI)-based regimens (69%) and most patients received peripheral blood stem cells (84%). Cyclosporine with methotrexate was the most common graft-versus-host-disease (GVHD) prophylaxis (54 %). In vivo T cell depletion was used in 44%. The 2-year overall survival (OS) was 69.4%, and leukemia -free survival (LFS) was 62.1%. The cumulative incidence of acute GVHD (aGVHD) grades II-IV at 100 days was 32.6%, and grades III-IV was 10%. The cumulative incidence of chronic GVHD (cGVHD) at 2 years was 37.3%, and extensive cGVHD was 16.8%. Multivariate analysis yielded significant associations between patient/transplant characteristics and outcomes. Advanced age at transplant was associated with poorer outcomes, including LFS (HR=1.11, p=0.004), GVHD-free, relapse-free survival (GRFS) (HR=1.06, p=0.04), OS (HR=1.12, p=0.002), and higher non-relapse mortality (NRM) (HR=1.23, p=0.001); HR for 10y increment. A later year of HSCT was associated with improved GFRS (HR=0.89, p=0.001), OS (HR=0.9, p=0.02), and decreased NRM (HR=0.82, p=0.008), aGVHD II-IV (HR=0.83, p=0.001), and cGVHD (HR=0.8, p=0.001); HR for 3y increment. TBI was specifically superior with improved LFS (HR=0.79, p=0.02), GRFS (HR=0.83, p=0.04), and lower relapse incidence (RI) (HR=0.65, p=0.001). Female-to-male transplant combination had a negative impact on GFRS (HR=1.21, p=0.02), OS (HR=1.23, p=0.048), and on the risk of cGVHD in general (HR=1.39, p=0.002) and specifically extensive cGVHD (HR=1.47, p=0.01). The use of in vivo T-cell depletion had significant benefits in terms of GFRS, aGVHD grade III-IV, cGVHD, and extensive cGVHD. Conclusion: This large study identified prognostic factors, such as age at transplant, donor type, and conditioning regimen, in influencing key outcomes including OS, LFS, GVHD incidence, and NRM in adult T-ALL patients undergoing allo-HSCT. Importantly, a significant improvement over time in post-transplant outcomes was noted. These findings hold great promise for new adapted treatment strategies and can serve as a benchmark for future studies in that setting.

Chromothripsis and Genomic Complexity As Pejorative Prognostic Markers in Pediatric and Adult T-ALL

Introduction: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplasm that accounts for 25% and 15% of adult and pediatric ALL, respectively. The prognosis is particularly poor in patients with relapsed disease, justifying the research for novel predictive markers of relapse.To date, a significant series of adult and pediatric T-ALL with analysis using high-resolution SNP-array is still missing. Methods: Here, we report a comprehensive high-resolution SNP-array profiling in a large well-characterized cohort of 317 T-ALL patients all uniformly treated according to the French FRALLE and GRAALL protocols. SNP-array results have been correlated with clinico-biological data including age range, immunophenotype, oncogenetic and survival data. Results: The SNP-array analysis detected at least one genomic imbalance (CNVs and UPDs combined) in 304/317 (96%) T-ALL cases with a mean of 6.3 (range 0-47) genomic imbalances per sample. Among genomic imbalances detected by SNP-array analysis, we identified 35 recurrent genomic imbalances (i.e. observed in at least 2% of T-ALL patients). The most common recurrent CNV was the del(9p) including CDKN2A/B at 9p21 (~70%). Unexpectedly, the second more frequent CNV was del(13q) minimally including RB1 and/or DLEU1 at 13q14 (~14%). The third more frequent CNV was the del(6q) minimally encompassing CASP8AP2 at 6q15 (~11%). The following recurrent CNVs were: del(12p) minimally including ETV6 and/or CDKN1B at 12p13 (~9%), del(18p) minimally including PTPN2 at 18p11 (~9%), del(1p) minimally including RPL22 at 1p36 (~9%), del(17q) minimally including NF1/SUZ12 at 17q11 (~8%). Two regions were identified as being recurrently affected by UPDs: 9p21 ( CDKN2A/B) (~28%), and 12q14 ( SH2B3) (~2%). Chromothripsis was detected in 6 (~2%) cases. The present study revealed relative minor differences in the genomic landscape of T-ALL according to age range consisting primarily of higher incidence of del(1p36)/ RPL22, del(13q14)/ RB1/DLEU1 del(12p13)/ ETV6/CDKN1B and del(18p11)/ PTPN2 and lower incidence of del(9p21) and UPD(9p21)/ CDKN2A/B in T-ALL older than 30 years. Comparison of SNP-array results according to immunophenotype and oncogenetic mainly highlighted the specific cytogenetic pattern of the SIL-TAL1 subgroup which exhibited a low genomic complexity with a lower number of genomic imbalances per sample and a lower diversity in genomic imbalances. Using the “survcutpoint” approach, we determined that a cutoff of 15 alterations was the most significant threshold to stratify the patients into a high- and a low-risk group of relapse according to the number of alterations detected with our SNP-array approach. Compared to patients harboring less than 15 alterations (n=296), patients with ≥ 15 genomic imbalances (n=21) had a significantly shorter event-free survival (EFS) and increased cumulative incidence of relapse (CIR)(4y-EFS: 38% vs. 64%; hazard ratio (HR): 2.1, 95%CI (1.2 - 3.7); P=.008, 4y-CIR: 53% vs. 26%; subdistribution hazard ratio (SHR): 2.9, 95%CI (1.5 - 5.4); P=.001) (Figure 1A). Additionally, survival analysis revealed the poor outcome conferred by the presence of chromothripsis (n=6) as well as gain(6q27)/ MLLT4 (n=10) and del(16p13)/ CREBBP (n=15) despite the low number of affected cases (4y-EFS: 17% vs. 63%; HR: 3.1, 95%CI (1.3 - 7.8); P=.01 and 4y-CIR: 83% vs. 26%; SHR: 5.0, 95%CI (2.2 - 11.3); P &amp;lt;.001 for chromothripsis; 4y-EFS: 30% vs. 63%; HR: 2.5, 95%CI (1.2 - 5.4); P=.002 and 4y-CIR: 57% vs. 27%; SHR: 2.8, 95%CI (0.96 - 8.1); P=.06 for gain (6q27)/ MLLT4; 4y-EFS: 40% vs. 63%; HR: 2.5, 95%CI (1.3 - 4.8); P=.005 and CIR in the FRALLE cohort; CIR: 4y-CIR: 50% vs. 21%; SHR: 5.4, 95%CI (1.5 - 20.0); P=.001 for del(16p13)/ CREBBP) (Figure 1B). Conclusion: Our study demonstrated that the whole genome analysis of imbalances provides new insights to refine the risk stratification in T-ALL. Notably, genomic complexity (≥ 15 genomic imbalances) and chromothripis demonstrated an association with an inferior outcome in terms of EFS and CIR in pediatric and adult T-ALL.

Anlotinib exerts an anti-T-cell acute lymphoblastic leukemia effect in vitro and in vivo

BackgroundDespite some progress having been made regarding the treatment of T-cell acute lymphoblastic leukemia (T-ALL), the prognosis of T-ALL, particularly adult T-ALL, is still poor. Identifying novel, effective anti-T-ALL drugs is of great significance. Anlotinib, an oral tyrosine kinase inhibitor currently utilized in the treatment of lung cancer, exhibited a promising anti-T-ALL effect. A comprehensive study should therefore be conducted to explore both the in vitro as well as in vivo mechanisms of the anti-T-ALL effects of anlotinib. MethodsCCK8 assays and flow cytometry were employed to investigate the viability, cell cycle distribution, and apoptosis of T-ALL cell lines when treated with anlotinib. T-ALL xenograft mouse models were established to examine the in vivo antileukemic effects of anlotinib. Cellular and molecular analysis of T-ALL were conducted to define the underlying mechanisms. ResultsIn vitro, anlotinib significantly inhibited the viability, induced G2/M phase arrest and apoptosis in T-ALL cell lines in a concentration-dependent pattern. In vivo, anlotinib also demonstrated a strong anti-tumor effect at doses that are well-tolerated. Interestingly, anlotinib could decrease the protein levels of the intracellular domains of NOTCH1 (ICN1) and c-Myc, two important targets for T-ALL. Mechanistically, anlotinib-induced c-Myc reduction was associated with proteasome-mediated degradation, while the ICN1 reduction was not due to protein degradation or transcriptional repression. ConclusionsThe present study showed that anlotinib may be a promising anti-T-ALL candidate drug, and simultaneous reduction of the protein levels of both ICN1 and c-Myc may contribute to the anti-T-ALL efficacy of anlotinib.

The Clinicogenomic Landscape of Induction Failure in Childhood and Young Adult T-Cell Acute Lymphoblastic Leukemia.

Failure to respond to induction chemotherapy portends a poor outcome in childhood acute lymphoblastic leukemia (ALL) and is more frequent in T-cell ALL (T-ALL) than B-cell ALL. We aimed to address the limited understanding of clinical and genetic factors that influence outcome in a cohort of patients with T-ALL induction failure (IF). We studied all cases of T-ALL IF on two consecutive multinational randomized trials, UKALL2003 and UKALL2011, to define risk factors, treatment, and outcomes. We performed multiomic profiling to characterize the genomic landscape. IF occurred in 10.3% of cases and was significantly associated with increasing age, occurring in 20% of patients age 16 years and older. Five-year overall survival (OS) rates were 52.1% in IF and 90.2% in responsive patients (P < .001). Despite increased use of nelarabine-based chemotherapy consolidated by hematopoietic stem-cell transplant in UKALL2011, there was no improvement in outcome. Persistent end-of-consolidation molecular residual disease resulted in a significantly worse outcome (5-year OS, 14.3% v 68.5%; HR, 4.10; 95% CI, 1.35 to 12.45; P = .0071). Genomic profiling revealed a heterogeneous picture with 25 different initiating lesions converging on 10 subtype-defining genes. There was a remarkable abundance of TAL1 noncoding lesions, associated with a dismal outcome (5-year OS, 12.5%). Combining TAL1 lesions with mutations in the MYC and RAS pathways produces a genetic stratifier that identifies patients highly likely to fail conventional therapy (5-year OS, 23.1% v 86.4%; HR, 6.84; 95% CI, 2.78 to 16.78; P < .0001) and who should therefore be considered for experimental agents. The outcome of IF in T-ALL remains poor with current therapy. The lack of a unifying genetic driver suggests alternative approaches, particularly using immunotherapy, are urgently needed.

Impact of T-cell Receptor Status on Mutational Landscape and Outcome in T-ALL.

Impact of T-cell Receptor Status on Mutational Landscape and Outcome in T-ALL.

Implication of Dynamin-2 (DNM2) Mutations in Adult T-cell Acute Lymphoblastic Leukemia.

The objective of the present study was to improve the risk stratification of T-cell Acute Lymphoblastic Leukemia (T-ALL) patients. It aimed to identify the frequency and clinical impact of DNM2 gene mutations among adult T-ALL cases. The current study included 25 T-ALL patients before starting their treatment. Mutational analysis of DNM2 gene (exons 18 and 22) was performed for all patients using Macrogen 3730 apparatus. We identified DNM2 gene mutations in 19 out of 25 (76%) patients. The detected mutations were either missense or deletion. Only active mutations (deletion) were associated with poor induction remission response and high frequency of relapse. Two novel mutations were addressed among the studied cohort of patients. They included c.1866G>C (p.V596L) and c.1872delA in exon 18. A high frequency of silent mutations was also found in T-ALL patients, but with no impact on clinical features. The DNM2 mutations were prevalent among adult T-ALL patients and might have a role in the pathogenesis of the disease. Active DNM2 mutations were associated with poor clinical outcome. Moreover, high frequency of DNM2 mutations indicated that these mutations could be utilized in detection of minimal residual disease in T-ALL patients.

Reverse Phase Protein Array Profiling Identifies Recurrent Protein Expression Patterns of DNA Damage-Related Proteins across Acute and Chronic Leukemia: Samples from Adults and the Children's Oncology Group.

DNA damage response (DNADR) recognition and repair (DDR) pathways affect carcinogenesis and therapy responsiveness in cancers, including leukemia. We measured protein expression levels of 16 DNADR and DDR proteins using the Reverse Phase Protein Array methodology in acute myeloid (AML) (n = 1310), T-cell acute lymphoblastic leukemia (T-ALL) (n = 361) and chronic lymphocytic leukemia (CLL) (n = 795) cases. Clustering analysis identified five protein expression clusters; three were unique compared to normal CD34+ cells. Individual protein expression differed by disease for 14/16 proteins, with five highest in CLL and nine in T-ALL, and by age in T-ALL and AML (six and eleven proteins, respectively), but not CLL (n = 0). Most (96%) of the CLL cases clustered in one cluster; the other 4% were characterized by higher frequencies of deletion 13q and 17p, and fared poorly (p < 0.001). T-ALL predominated in C1 and AML in C5, but both occurred in all four acute-dominated clusters. Protein clusters showed similar implications for survival and remission duration in pediatric and adult T-ALL and AML populations, with C5 doing best in all. In summary, DNADR and DDR protein expression was abnormal in leukemia and formed recurrent clusters that were shared across the leukemias with shared prognostic implications across diseases, and individual proteins showed age- and disease-related differences.

The oncogenetic landscape and clinical impact of BCL11B alterations in adult and pediatric T-cell acute lymphoblastic leukemia.

Not available.

FAT1 expression in T-cell acute lymphoblastic leukemia (T-ALL) modulates proliferation and WNT signaling

FAT atypical cadherin 1 (FAT1), a transmembrane protein, is frequently mutated in various cancer types and has been described as context-dependent tumor suppressor or oncogene. The FAT1 gene is mutated in 12–16% of T-cell acute leukemia (T-ALL) and aberrantly expressed in about 54% of T-ALL cases contrasted with absent expression in normal T-cells. Here, we characterized FAT1 expression and profiled the methylation status from T-ALL patients. In our T-ALL cohort, 53% of patient samples were FAT1 positive (FAT1pos) compared to only 16% FAT1 positivity in early T-ALL patient samples. Aberrant expression of FAT1 was strongly associated with FAT1 promotor hypomethylation, yet a subset, mainly consisting of TLX1-driven T-ALL patient samples showed methylation-independent high FAT1 expression. Genes correlating with FAT1 expression revealed enrichment in WNT signaling genes representing the most enriched single pathway. FAT1 knockdown or knockout led to impaired proliferation and downregulation of WNT pathway target genes (CCND1, MYC, LEF1), while FAT1 overexpressing conveyed a proliferative advantage. To conclude, we characterized a subtype pattern of FAT1 gene expression in adult T-ALL patients correlating with promotor methylation status. FAT1 dependent proliferation and WNT signaling discloses an impact on deeper understanding of T-ALL leukemogenesis as a fundament for prospective therapeutic strategies.

DNMT3A Mutations Enhance T-ALL Survival, Which Is Reversed By JAK/STAT Inhibition

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplasm of T-cell progenitors, which accounts for approximately 15% and 25% of pediatric and adult acute lymphoblastic leukemia (ALL) cases respectively. In T-ALL, accumulation of genomic abnormalities often leads to the aberrant expression of select groups of transcription factors that result in increased proliferation, cell survival and impaired differentiation of T-cell progenitors, which give rise to this disease The Notch signaling pathway is essential for T-lymphopoiesis and T-cell differentiation from thymic precursors. Moreover, gain-of-function mutations in this pathway act as the major genetic lesions that drive T-ALL development (over 60% of adult cases). The de novo DNA methyltransferase enzyme (DNMT3A) is one of the three genes in mammals encoding for enzymes with DNA methyltransferase activity and mutations in this gene are frequently associated with NOTCH1 mutations in adult T-ALL. Interestingly, DNMT3A mutations are rare in pediatric and adolescent T-ALL. Prognosis of pediatric and adolescent T-ALL cases has seen major improvements during recent years with the introduction of high-dose, multi-agent chemotherapy regimens which has not carried over to adult T-ALL patients. These factors suggest the involvement of genetic differences between adult and pediatric T-ALL patients for the differential clinical outcomes Previous work in our lab has shown that mouse T-ALL cells genetically deficient for Dnmt3a are less apoptotic and have increased JAK/STAT signaling. Here, we demonstrate that DNMT3A-mutant human T-ALL patient derived xenografts (PDXs) are resistant to the chemotherapeutic drugs dexamethasone (DEX) and doxorubicin compared to T-ALL patient specimens with wild-type DNMT3A. However, when primary DNMT3A-mutant human T-ALL cells were treated with dexamethasone or doxorubicin in combination with the JAK/STAT inhibitor ruxolitinib (RUX), we observed the survival benefit of DNMT3A-mutant T-ALL cells were mitigated, suggesting enhanced JAK/STAT signaling promotes survival in T-ALL patient cells with DNMT3A mutations. To investigate the genes and pathways that endows DNMT3A-mutant T-ALL cells chemoresistance, gene expression profiling was performed by RNA-seq on DNMT3A-mutant, and wild-type human T-ALL cells treated with DEX+RUX. Gene set enrichment analysis (GSEA) showed that MTORC1, IL2-STAT5, E2F and G2M pathways were significantly downregulated in DEX+RUX treated DNMT3A-mutant PDX samples. E2F and G2M pathways are associated with cell cycle progression and further analysis of these two pathways revealed BIRC5 as a common gene that is downregulated in both these pathways following DEX+RUX treatment. Furthermore, our RNA-seq analysis demonstrated that the expression of BIRC5 in DNMT3A-mutant PDXs is downregulated upon DEX+RUX treatment compared to the DNMT3A-wildtype group. Moreover, baseline BIRC5 expression is higher in DNMT3A-mutant T-ALL PDXs compared to the DNMT3A-wildtype group. Following treatment with the BIRC5 inhibitor YM155, the viability of DNMT3A-mutant T-ALL cells decrease significantly compared to DNMT3A-wildtype PDXs, which were relatively unaffected. In addition, we observed a similar reduction in cell viability in DNMT3A-mutant PDXs following BIRC5 siRNA nucleofection compared to the DNMT3A-wildtype group. Cumulatively, these studies reveal that enhanced JAK/STAT signaling promotes survival and chemoresistance in T-ALL patient cells with DNMT3A mutations and identify the JAK/STAT target gene BIRC5 as a specific genetic dependency of DNMT3A-mutant T-ALL cells. These data provide a critical first step towards a novel target for precision medicine approaches for this patient group.

Conditional Knock-out of Phf6 Decreases CD44 Expression on Naïve CD4+ T Cells and Skews the Differentiation Toward Regulatory T Cells in Mice

Human PHF6 (plant homeodomain (PHD) finger 6), located in X chromosome, is highly expressed in the central nervous system as well as B- and T-lymphoid cells in human and mice, and involved in multiple physiological pathways through chromatin regulation by interaction with the nucleosome remodeling and deacetylation (NuRD) complex. Germline mutations of PHF6 lead to Börjeson-Forssman-Lehmann syndrome (BFLS), a rare X-linked disorder with distinctive facial features and mental retardation. Meanwhile, somatic PHF6 mutations are present in 11% to 39.5% of adult T-cell acute lymphoblastic leukemia (T-ALL) patients. PHF6 functions as a tumor suppressor gene in T-ALL. Conditional knock-out (cKO) of Phf6 in hematopoietic cells in mice enhances self-renewal potential of hematopoietic stem cells and accelerates the oncogenesis of ICN1 overexpression-induced T-ALL. However, the roles of Phf6 in regulating the development and function of lymphoid cells are not explored yet. In our previous study (YC Hsu et al.Blood adv. 2019), we have shown decreased circulating CD4+ T cells in the Phf6 cKO (Phf6 cKO, Phf6F/F or Phf6F/Y;Vav1-Cre) mice compared with the wild type (WT, Phf6F/F or Phf6F/Y) mice. While the percentages of naïve, effector, and effector memory CD4+ T cells, defined by surface CD44 and CD62L, were comparable between the WT and Phf6 cKO mice, we found significantly lower expression of surface CD44 on naïve CD4+ T cells in the Phf6 cKO mice. To assess the effects of Phf6 on the development of the T cells, we performed bulk RNA sequencing (RNAseq) of thymocytes from 3 pairs of mice. GSEA for these transcriptomes revealed enrichment of the interferon γ, interferon α, and FOXP3 targets. These results support our hypothesis that Phf6 has a role in T cell function and regulation. Consisted with the RNAseq data showing enrichment of the FOXP3 targets in the thymocytes of the Phf6 cKO mice, we discovered that the pooled Treg cells in steady state, containing a majority of natural Treg cells (nTreg cells) and some induced Treg cells (iTreg cells), were significantly increased in the peripheral blood and lymph nodes of the Phf6 cKO mice as revealed by the intracellular Foxp3. (Figure 1a) We further investigated if Phf6 has a role in iTreg cells. To this end, we performed in vitro simulation on splenocytes of the WT and Phf6 cKO mice with CD3 mAB, CD28 mAB, hTGFb1 and mIL2 and found the CD4+Phf6 cKO splenocytes were more prone to differentiate into the CD25+Foxp3+ Treg cells compared with that of the WT mice. (Figure 1b) Collectively, conditional knock-out of Phf6 in mice can skew lymphocyte differentiation toward Treg cells not only in steady state, but also under in vitro regulatory T cell priming condition. To further explore the possible mechanisms, we analyzed single cell RNAseq of splenic T cells of WT and Phf6 cKO mice at steady state and cataloging 7 populations as defined by distinct transcriptomic patterns, including 5 for CD4+ T cells and 2 for CD8+ T cells. (Figure 1c) Compared with the WT, the T cells of Phf6 cKO mice had a larger proportion of Treg cells, which significantly expressed higher Fyb and Eno1. (Figure 1d, e and f) According to the literature, FYB, also known as ADAP (Adhesion and degranulation promoting adaptor protein), is a positive regulator of T-cell development, TCR/CD3-induced T-cell activation, and IL-2 and IFN-g production. (E K Griffiths et al. Science. 2001, Zhang Y et al. Immunology. 2012) Meanwhile, the glycolytic enzyme enolase-1, encoded by Eno1, can stimulate glycolysis, on which the induction and function of induced Treg cells highly depend. (V De Rosa et al. Nat Immunol. 2015) In conclusion, we show that Phf6 is a novel regulator of Treg cells perhaps through ADAP and glycolysis pathways. We will further explore the functions and mechanisms of the Phf6 on T cell regulation. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

RPPA-Profiling in Pediatric and Adult T-Cell Acute Lymphoblastic Leukemia Identifies Protein Patterns Associated with Outcome

RPPA-Profiling in Pediatric and Adult T-Cell Acute Lymphoblastic Leukemia Identifies Protein Patterns Associated with Outcome