T-cell Lymphoblastic Lymphoma Patients Research Articles

NOTCH1 fusions in pediatric T-cell lymphoblastic lymphoma: Ahigh-risk subgroup with CCL17 (TARC) levels as diagnostic biomarker.

Twenty percent of children with T-cell lymphoblastic lymphoma (T-LBL) will relapse and have an extremely poor outcome. Currently, we can identify a genetically low-risk subgroup in pediatric T-LBL, yet these high-risk patients who need intensified or alternative treatment options remain undetected. Therefore, there is an urgent need to recognize these high-risk T-LBL patients through identification of molecular characteristics and biomarkers. By using RNA sequencing which was performed in 29/49 T-LBL patients who were diagnosed in the Princess Maxima Center for Pediatric Oncology between 2018 and 2023, we discovered a previously unknown high-risk biological subgroup of children with T-LBL. This subgroup is characterized by NOTCH1 gene fusions, found in 21% of our T-LBL cohort (6/29). All patients presented with a large mediastinal mass, pleural/pericardial effusions, and absence of blasts in the bone marrow, blood, and central nervous system. Blood CCL17 (C-C Motif Chemokine Ligand 17, TARC) levels were measured at diagnosis in 26/29 patients, and all six patients with NOTCH1 gene fusions patients exclusively expressed highly elevated blood CCL17 levels, defining a novel and previously not known clinically relevant biomarker for T-cell lymphoblastic lymphoma. Four out of these six patients relapsed during therapy, a fifth developed a therapy-related acute myeloid leukemia during maintenance therapy. These data indicate that T-LBL patients with a NOTCH1 fusion have a high risk of relapse which can be easily identified using a blood CCL17 screening at diagnosis. Further molecular characterization through NOTCH1 gene fusion analysis offers these patients the opportunity for treatment intensification or new treatment strategies.

Differential impact of asparaginase discontinuation on outcomes of children with T-cell acute lymphoblastic leukemia and T-cell lymphoblastic lymphoma.

Asparaginase is essential for treating T-cell acute lymphoblastic leukemia (T-ALL). Despite the ongoing debate on whether T-ALL and T-cell lymphoblastic lymphoma (T-LBL) are the same disease entity or two distinct diseases, patients with T-LBL often receive the same or similar treatment protocols as those with T-ALL. The outcomes of patients with or without L-asparaginase discontinuation were retrospectively analyzed among four national protocols: Japan Association of Childhood Leukemia Study (JACLS) ALL-02 and ALL-97 for T-ALL and Japanese Pediatric Leukemia/Lymphoma Study Group ALB-NHL03 and JACLS NHL-98 for T-LBL. The hazard ratio (HR) was calculated with the Cox regression model by considering L-asparaginase discontinuation as a time-dependent variable. In total, 199 patients with T-ALL, and 133 patients with T-LBL were included. L-asparaginase discontinuation compromised event-free survival (EFS) of T-ALL patients (ALL-02: HR 3.32, 95% confidence interval [CI] 1.40-7.90; ALL-97: HR 3.39, 95%CI 1.19-9.67). Conversely, EFS compromise was not detected among T-LBL patients (ALB-NHL03: HR 1.39, 95%CI 0.41-4.68; NHL-98: HR 0.92, 95%CI 0.11-7.60). The effects of L-asparaginase discontinuation differed between T-ALL and T-LBL. We assume that the differential impact results from (1) the inherent differential response to L-asparaginase between them and/or (2) a less stringent assessment of early treatment response in T-LBL than in T-ALL. Given the poor salvage rate of refractory or relapsed T-ALL and T-LBL, optimization of the frontline therapy is critical, and the current study provides a new suggestion for further treatment modifications. However, larger studies in contemporary intensified treatment protocols are required.

Key candidate genes and pathways in T lymphoblastic leukemia/lymphoma identified by bioinformatics and serological analyses.

T-cell acute lymphoblastic leukemia (T-ALL)/T-cell lymphoblastic lymphoma (T-LBL) is an uncommon but highly aggressive hematological malignancy. It has high recurrence and mortality rates and is challenging to treat. This study conducted bioinformatics analyses, compared genetic expression profiles of healthy controls with patients having T-ALL/T-LBL, and verified the results through serological indicators. Data were acquired from the GSE48558 dataset from Gene Expression Omnibus (GEO). T-ALL patients and normal T cells-related differentially expressed genes (DEGs) were investigated using the online analysis tool GEO2R in GEO, identifying 78 upregulated and 130 downregulated genes. Gene Ontology (GO) and protein-protein interaction (PPI) network analyses of the top 10 DEGs showed enrichment in pathways linked to abnormal mitotic cell cycles, chromosomal instability, dysfunction of inflammatory mediators, and functional defects in T-cells, natural killer (NK) cells, and immune checkpoints. The DEGs were then validated by examining blood indices in samples obtained from patients, comparing the T-ALL/T-LBL group with the control group. Significant differences were observed in the levels of various blood components between T-ALL and T-LBL patients. These components include neutrophils, lymphocyte percentage, hemoglobin (HGB), total protein, globulin, erythropoietin (EPO) levels, thrombin time (TT), D-dimer (DD), and C-reactive protein (CRP). Additionally, there were significant differences in peripheral blood leukocyte count, absolute lymphocyte count, creatinine, cholesterol, low-density lipoprotein, folate, and thrombin times. The genes and pathways associated with T-LBL/T-ALL were identified, and peripheral blood HGB, EPO, TT, DD, and CRP were key molecular markers. This will assist the diagnosis of T-ALL/T-LBL, with applications for differential diagnosis, treatment, and prognosis.

Targeting P21-activated kinase suppresses proliferation and enhances chemosensitivity in T-cell lymphoblastic lymphoma.

T-cell lymphoblastic lymphoma (T-LBL) is a highly aggressive non-Hodgkin lymphoma with a poor prognosis. P21-activated kinase (PAK) is a component of the gene expression-based classifier that can predict the prognosis of T-LBL. However, the role of PAK in T-LBL progression and survival remains poorly understood. Herein, we found that the expression of PAK1 was significantly higher in T-LBL cell lines (Jurkat, SUP-T1, and CCRF-CEM) compared to the human T-lymphoid cell line. Moreover, PAK2 mRNA level of 32 relapsed T-LBL patients was significantly higher than that of 37 cases without relapse (P = .012). T-LBL patients with high PAK1 and PAK2 expression had significantly shorter median RFS than those with low PAK1 and PAK2 expression (PAK1, P = .028; PAK2, P = .027; PAK1/2, P = .032). PAK inhibitors, PF3758309 (PF) and FRAX597, could suppress the proliferation of T-LBL cells by blocking the G1/S cell cycle phase transition. Besides, PF could enhance the chemosensitivity to doxorubicin in vitro and in vivo. Mechanistically, through western blotting and RNA sequencing, we identified that PF could inhibit the phosphorylation of PAK1/2 and downregulate the expression of cyclin D1, NF-κB and cell adhesion signaling pathways in T-LBL cell lines. These findings suggest that PAK might be associated with T-LBL recurrence and further found that PAK inhibitors could suppress proliferation and enhance chemosensitivity of T-LBL cells treated with doxorubicin. Collectively, our present study underscores the potential therapeutic effect of inhibiting PAK in T-LBL therapy.

17th International Conference on Malignant Lymphoma, Palazzo dei Congressi, Lugano, Switzerland, 13 - 17 June, 2023.

Introduction: Mature B-cell non-Hodgkin's lymphoma (MB-NHL) and T-cell lymphoblastic lymphoma (T-LBL) account for ∼50% and ∼20%, respectively, among the different pediatric NHL entities. Our understanding of the genetic lesions and profiling of these patients may help to improve the clinical management of patients with these lymphomas. The aim of this study is to analyze the mutational status and the associations between genetic features and treatment outcomes in the Chinese pediatric lymphoma cohort. Methods: A total of 207 patients treated at multiple clinical centers of the Chinese Children's Lymphoma Collaborative Group (CNCL) from 2017 to 2023 were included in this retrospective study. Targeted next-generation sequencing (t-NGS) with a panel of lymphoma-related genes was performed on tumor samples collected at the time of initial diagnosis and at tumor of refractory or relapse (r/r), and the correlations of somatic mutations with survival rates as well as with relapse and other clinical factors were analyzed. Results: A total of 136 driver genes with somatic mutations were detected in the entire cohort. In 133 pediatric MB-NHL patients, the most frequently mutated genes from 77 initially diagnostic samples were ID3 (52%), TP53 (47%), CCND3 (30%), ARID1A (29%), and DDX3X (27%) (Figure 1a). In 56 r/r samples, the most commonly mutated genes were TP53 (84%), followed by ID3 (59%), ARID1A (41%), CCND3 (32%), and DDX3X (25%) (Figure 1b). TP53 mutation was significantly more frequent in r/r samples than that in initially diagnostic samples (p < 0.001) and patients with TP53 mutations had poor survival (log-rank p = 0.0013, Figure 1c). Among patients treated with chimeric antigen receptor T-cell (CAR-T) therapy, those with ARID1A mutations exhibited poorer response to CAR-T treatment and shorter overall survival compared with the patients without such mutations (mOS = 180 days, log-rank p = 0.00081, Figure 1d). In 74 pediatric T-LBL patients,the most commonly mutated genes were NOTCH1 (50%), followed by FBXW7 (26%), JAK1 (16%), NRAS (16%), and JAK3 (11%) (Figure 1e). Genes in the JAK signaling pathway were more frequently mutated in Chinese patients than the corresponding western patients, such as JAK1 (16% vs. 2%) and JAK3 (11% vs. 5%). Further analysis showed that the incidence of NOTCH1 (68% vs. 27%) and FBXW7 (37% vs. 12%) mutations in patient group with initial remission (n = 41) was significantly higher than that in patient group with r/r T-LBL (n = 33) (p = 0.005, p = 0.036, respectively, Figure 1f), indicating that NOTCH1 and FBXW7mutations were associated with good prognosis. Keywords: diagnostic and prognostic biomarkers, immunotherapy, non-Hodgkin (pediatric, adolescent, and young adult) No conflicts of interests pertinent to the abstract.

The β-catenin-LINC00183-miR-371b-5p-Smad2/LEF1 axis promotes adult T-cell lymphoblastic lymphoma progression and chemoresistance

BackgroundHigh-intensity chemotherapy regimens are often used in adult T-cell lymphoblastic lymphoma (T-LBL) patients. Nevertheless, the response rate remains unsatisfactory due to emergence of chemoresistance. Growing evidence has shown that long non-coding RNAs (lncRNAs) are involved in tumor progression and chemoresistance. Herein, we investigated the potential role of lncRNAs in T-LBLs.MethodsRNAseq was used to screen and identify candidate lncRNAs associated with T-LBL progression and chemoresistance. Luciferase reporter assay was used to examine the binding of miR-371b-5p to the 3’UTR of Smad2 and LEF1, and the binding of TCF-4/LEF1 to the promoter of LINC00183. Chromatin immunoprecipitation assay was undertaken to analyze the connection between LEF1 and the LINC00183 promoter region. RNA immunoprecipitation assays were used to explore the mechanism whereby LINC00183 regulated miR-371b-5p. MTT and flow cytometry assays were used to measure apoptosis of T-LBL cells.ResultsLINC00183 was upregulated in T-LBL progression and chemoresistant tissues in both the Sun Yat-sen University Cancer Center dataset and the First Affiliated Hospital of Anhui Medical University dataset. High expression of LINC00183 was correlated with poorer overall survival and progression-free survival of T-LBL patients compared to those with low expression of LINC00183. Furthermore, miR-371b-5p was negatively regulated by LINC00183. In vivo and in vitro assays showed that LINC00183-mediated T-LBL chemoresistance depended on miR-371b-5p expression. The direct binding of miR-371b-5p to Smad2 and LEF1 was verified by luciferase assays. It was shown that TCF4/LEF1 could bind to the LINC00183 promoter site and increase its transcript level. Downregulation of miR-371b-5p led to increased expression of Smad2/LEF1, and in turn increased LINC00183 expression. Additionally, phospho-Smad2 promotes nuclear translocation of β-catenin, LINC00183 downregulation decreased chemoresistance induced by β-catenin and TGF-β1 in T-LBL cells.ConclusionWe unraveled a β-catenin-LINC00183-miR-371b-5p-Smad2/LEF1 feedback loop that promotes T-LBL progression and chemoresistance, indicating that LINC00183 may serve as a potential therapeutic target in T-LBLs.

Clinical and prognostic role of 2-[18F]FDG PET/CT and sarcopenia in treatment-naïve patients with T-cell lymphoblastic lymphoma.

T-cell lymphoblastic lymphoma (T-LBL) is a rare and highly aggressive non-Hodgkin lymphoma. This study aimed to explore the role of 2-[18F]FDG PET/CT, sarcopenia, clinical features, and treatment regimens in 49 treatment-naïve patients with T-LBL, and assess their predictive value in the prognosis. Sarcopenia was measured as skeletal muscle index (SMI) at L3 level from the CT component of PET/CT images. All 49 patients (35 males, 14 females; median age, 26years [range, 3-66years]) were enrolled in this study, including 36 adult patients and 13 pediatric patients. Lymph nodes, thymus, bone marrow, and pleura were the most common involved sites of T-LBL. The median SUVmax, MTV, and TLG of all lesions in these 49 patients were 12.4 (range, 4.2-40.5), 532.6 (17.4-3518.1), and 2112.2 (53.9-18,699.2), respectively. Eighteen out of 49 patients (36.7%) were diagnosed with sarcopenia. Sarcopenia patients had lower BMI and SUVmax of muscle at L3 level than non-sarcopenia patients (P < 0.05). Univariate Cox regression analysis indicated that higher MTV and TLG and intrathecal therapy (IT) were associated with longer progression-free survival (PFS) and overall survival (OS), while multivariate Cox regression analysis showed that TLG and IT were independent predictors for PFS, and only IT was an independent predictor for OS. In conclusion, low BMI and SUVmax of muscle at L3 level correlated with sarcopenia in T-LBL patients. Higher initial MTV and TLG and receiving IT were associated with better prognosis in T-LBL patients. TLG and IT, but not sarcopenia, were independent prognostic factors.

Diverse mutations and structural variations contribute to Notch signaling deregulation in paediatric T-cell lymphoblastic lymphoma.

T-cell lymphoblastic lymphoma (T-LBL) is an aggressive neoplasm closely related to T-cell acute lymphoblastic leukaemia (T-ALL). Despite their similarities, and contrary to T-ALL, studies on paediatric T-LBL are scarce and, therefore, its molecular landscape has not yet been fully elucidated. Thus, the aims of this study were to characterize the genetic and molecular heterogeneity of paediatric T-LBL and to evaluate novel molecular markers differentiating this entity from T-ALL. Thirty-three paediatric T-LBL patients were analyzed using an integrated approach, including targeted next-generation sequencing, RNA-sequencing transcriptome analysis and copy-number arrays. Copy number and mutational analyses allowed the detection of recurrent homozygous deletions of 9p/CDKN2A (78%), trisomy 20 (19%) and gains of 17q24-q25 (16%), as well as frequent mutations of NOTCH1 (62%), followed by the BCL11B (23%), WT1 (19%) and FBXW7, PHF6 and RPL10 genes (15%, respectively). This genetic profile did not differ from that described in T-ALL in terms of mutation incidence and global genomic complexity level, but unveiled virtually exclusive 17q25 gains and trisomy 20 in T-LBL. Additionally, we identified novel gene fusions in paediatric T-LBL, including NOTCH1-IKZF2, RNGTT-SNAP91 and DDX3X-MLLT10, the last being the only one previously described in T-ALL. Moreover, clinical correlations highlighted the presence of Notch pathway alterations as a factor related to favourable outcome. In summary, the genomic landscape of paediatric T-LBL is similar to that observed in T-ALL, and Notch signaling pathway deregulation remains the cornerstone in its pathogenesis, including not only mutations but fusion genes targeting NOTCH1.

Comprehensive view on genetic features, therapeutic modalities and prognostic models in adult T-cell lymphoblastic lymphoma.

Adult T-cell lymphoblastic lymphoma (T-LBL) is a rare and aggressive subtype of non-Hodgkin's lymphoma that differs from pediatric T-LBL and has a worse prognosis. Due to its rarity, little is known about the genetic and molecular characteristics, optimal treatment modalities, and prognostic factors of adult T-LBL. Therefore, we summarized the existing studies to comprehensively discuss the above issues in this review. Genetic mutations of NOTCH1/FBXW7, PTEN, RAS, and KMT2D, together with abnormal activation of signaling pathways, such as the JAK-STAT signaling pathway were described. We also discussed the therapeutic modalities. Once diagnosed, adult T-LBL patients should receive intensive or pediatric acute lymphoblastic leukemia regimen and central nervous system prophylaxis as soon as possible, and cranial radiation-free protocols are appropriate. Mediastinal radiotherapy improves clinical outcomes, but adverse events are of concern. Hematopoietic stem cell transplantation may be considered for adult T-LBL patients with high-risk factors or those with relapsed/refractory disease. Besides, several novel prognostic models have been constructed, such as the 5-miRNAs-based classifier, 11-gene-based classifier, and 4-CpG-based classifier, which have presented significant prognostic value in adult T-LBL.

Central nervous system neurotoxicity associated with nelarabine in T-cell acute lymphoblastic leukemia.

Nelarabine, a prodrug of arabinosylguanine has lineage-specific toxicity for T lymphoblasts and is used to treat refractory or relapsed T-cell acute lymphoblastic leukemia and T-cell lymphoblastic lymphoma patients. The most commonly observed adverse effects associated with nelarabine are mainly hematological, i.e. neutropenia, anemia, and thrombocytopenia. Additionally, neurological, and gastrointestinal toxicities have been reported. Central nervous system neurotoxicity associated with nelarabine is very rare. A 37-year-old man patient diagnosed with T-cell acute lymphoblastic leukemia had experienced generalized tonic-clonic seizure which lasted for a few seconds and upper extremity weakness after three weeks of the nelarabine infusion. Computed tomography and magnetic resonance imaging have shown periventricular and nucleus caudatus abnormalities. Radiological findings suggested toxic leukoencephalopathy and acute infarct of right nucleus caudatus. After high-dose steroids, intravenous immunoglobulin, and support treatment, his neurologic symptoms disappeared except for mild peroral numbness. However, radiological sequelae persisted despite clinical improvement. Physicians involved in the care of these patients who use nelarabine should be aware of the fact that cerebral toxicity of the nelarabine may occur especially in the presence of predisposing factors. It is crucial to monitor closely those patients receiving nelarabine and also those who have additional predisposing factors for neurotoxicity.

Survival and prognostic analysis of T-cell lymphoblastic lymphoma patients treated with dose-adjusted BFM-90 regimen.

We aimed to investigate the long-term prognosis and prognostic factors of T-cell lymphoblastic lymphoma (T-LBL) patients who received dose-adjusted Berlin–Frankfurt–Münster (BFM)-90 regimen as first-line therapy in our center. A total of 145 T-LBL patients who underwent first-line dose-adjusted BFM-90 was retrospectively reviewed. Conditional survival analysis was used to evaluate the long-term prognosis of patients. Receiver operating characteristic (ROC) curve was applied to determine the optimal cut-off value for neutrophil-to-lymphocyte ratio (NLR). Estimated 3-year overall survival (OS) and progression-free survival (PFS) rates for overall were 66.8% and 58.4%, respectively. Conditional survival analysis showed that for patients having survived 3 and 5 years or more after the completion of the treatment, the estimated subsequent 3-year OS thereafter increased to 85.7% and 94.3, respectively. Patients receiving consolidation APBSCT (Autologous peripheral blood stem cell transplantation) after BFM-90 regimen had superior 3-year OS than those with non-APBSCT (79.1% vs. 33.4%, p<0.001). We also discovered that baseline NLR ≥4.95 was negatively associated with OS (HR=2.75, 95% CI 1.55-4.89, p=0.015) and PFS (HR=2.07, 95% CI 1.25-4.96, p=0.021) via multivariable analysis. Conclusions: The survival probability of T-LBL patients treated with first-line dose-adjusted BFM-90 has improved significantly as patients have survived for every additional year. The addition of consolidation APBSCT following dose-adjusted BFM-90 therapy bring further survival benefits for those patients. Baseline NLR ≥4.95 was an independent risk factor for T-LBL patients in our study.

High Prevalence of Constitutional Mismatch Repair Deficiency in a Pediatric T-cell Lymphoblastic Lymphoma Cohort.

This study describes the clinical characteristics of a complete Dutch T-cell lymphoblastic lymphoma (T-LBL) cohort, including second primary malignancies and comorbidities. We show that over 10% of patients in this complete T-LBL cohort have been diagnosed with a cancer predisposition syndrome (CPS), consisting almost exclusively of constitutional mismatch repair deficiency (CMMRD). The clinical characteristics of sporadic T-LBL patients were compared with T-LBL patients that have been diagnosed with CMMRD. This shows that disease presentation is comparable but that disease localization in CMMRD patients might be more localized. The percentage of CPS seems reliable considering the completeness of the cohort of Dutch T-LBL patients and might even be an underestimation (possibility of undiagnosed CPS patients in cohort). As the frequency of an underlying predisposition syndrome among T-LBL patients may be underestimated at present, we advocate for screening all pediatric T-LBL patients for the presence of germline mutations in mismatch repair genes.

MicroRNA-211 attenuates cell proliferation in T-cell lymphoblastic lymphoma through targeting TCF12

T-cell lymphoblastic lymphoma (T-LBL) is a class of highly aggressive hematologic neoplasms originating from progenitor T-lymphocytes. MicroRNA (miRNA) is an endogenous RNA molecule with 22 nucleotides in length. Accumulated evidence suggests that miRNA functions as a key regulator in human cancer. Herein, by in silico analysis, we found that miR-211 was a decreased miRNA in T-LBL in high-throughput sequencing data, which was subsequently verified in our cohort. Low miR-211 was closely correlated with bulky disease, high ann arbor stage, relapse and poor prognosis. miR-211 was regulated by N6-methyladenosine (m6A) modification, specifically, m6A methyltransferase METTL14 methylated primary miR-211 (pri-miR-211), expediting pri-miR-211 processing via recruiting DGCR8. Functionally, miR-211 overexpression significantly reduced T-LBL cell viability, DNA synthesis rate and spheroid formation ability, whereas silencing of miR-211 had the opposite effects. In addition, we established the xenograft tumor model and found that miR-211 remarkably inhibited tumor growth in vivo. Further, TCF12 was the direct target of miR-211, miR-211 bound to TCF12 mRNA 3`-untranslated region (UTR) and increased its decay, overexpression of TCF12 could effectively rescue the weakened malignant behavior of T-LBL cells caused by miR-211 overexpression. Collectively, our data clearly demonstrate that miR-211 is a novel tumor suppressor in T-LBL, targeting of miR-211/TCF12 axis may be a potential treatment for T-LBL patients.

Outcome of Allogeneic Stem Cell Transplantation in T Cell Lymphoblastic Lymphoma

Background T cell lymphoblastic lymphoma (T-LBL) is an aggressive form of non-Hodgkin lymphoma occurring in predominantly adolescent and young adults.Although T-LBL is achemotherapy-sensitive disease, relapse is common with conventional chemotherapies. Allogeneic stem cell transplantation (allo-SCT) may be effective in patients with high-risk features or relapsed/refractory patients.Objective To investigate the effect of allo-SCT in the treatment of T-LBL.Methods T-LBL patients who received allo-SCT from August 2007 to March 2017 in our center were recruited in this retrospective study.Results (1) 37 T-LBL patients were enrolled in this study, including 25 males and 12 females with a median age of 21 years old (ranged from 8 to 47 years old). A total of 10 (27%) from HLA-matched siblings,24 (65%) patients received grafts from haploidentical family donors, 2 (5%) from matched unrelated donors,and 1(3%) from cord blood. (2) With a median follow up of 20 months (4 - 119 months), 3 year overall survival (OS) ,progression-free survival (PFS) and relapse were 71.7%, 69.5% and 23.6%,respectively. (3) 12/37 patients developed acute graft-versus-host diease (GVHD) ,including 6 (16%) with grade III-IV aGVHD. 8/37 patients developed chronic GVHD. The transplant-related mortality was 13.5%.(4) Allo-SCT in first complete remission (CR1) was associated with improved OS (3-year estimates: 79.1% vs 45.5%, P=0.001) and improved PFS (62.5% vs 39.1%, P=0.001). On multivariable analysis, allo-SCT in CR1 was the factor associated with the prolonged OS (hazard ratio [HR] = 0.095;95% confidence interval [CI] : 0.014-0.655; P=0.017) and PFS (HR=,0.085 95% CI=0.014-0.515; P=0.007).Conclusion Allo-SCT for T-LBL patients revealed a proming outcome. Disease status the only independent factor affecting the OS and PFS, and allo-SCT should be recommended in CR1. DisclosuresNo relevant conflicts of interest to declare.

Prognostic Value of the Immunological Subtypes of Adolescent and Adult T-Cell Lymphoblastic Lymphoma; an Ultra-High-Risk Pro-T/CD2(-) Subtype.

Simple SummaryT-cell lymphoblastic lymphoma (T-LBL) is extremely rare and aggressive with no practical risk model defined. Considering the controversies over the prognostic value of T-LBL immunological subtypes, we re-evaluated 49 subsequent adult T-LBL patients treated according to the German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia protocols, with 85.7% with complete remissions. To the best of our knowledge, this is the largest study of T-LBL diagnosed by flow-cytometry of the material obtained by fine-needle aspiration biopsy. We show that (1) CD2 status and age are powerful independent prognostic factors influencing overall survival and the risk of treatment failure; (2) the early/pro-T/CD2(−) subtype is associated with extremely poor outcomes; and (3) poor outcomes in ETP vs. non-ETP are strikingly consistent with the pro-T CD2(−) subtype. The lack of CD2 expression in T-LBL emerges as a new marker of an ultra-high-risk of treatment failure. We show here that ETP is a non-uniform entity, where the outcome depends on the CD2 status.(1) Background: T-cell lymphoblastic lymphoma (T-LBL) is extremely rare and highly aggressive, with no practical risk model defined yet. The prognostic value of T-LBL immunological subtypes is still a matter of controversy. (2) Methods: We re-evaluated 49 subsequent adult T-LBL patients treated according to the German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia (GMALL) protocols, 05/93 (n = 20) and T-LBL 1/2004 (n = 29), 85.7% of which achieved complete remission (CR). (3) Results: The 5/10-year overall survival (OS) and event-free survival (EFS) were 62%/59% and 48%/43%, respectively. In 96% of patients, flow cytometry analyses defining the WHO 2008 immunophenotypes were available. Cortical, early/pro-T/CD2(−), early/pre-T/CD2(+), and mature subtypes were identified in 59.5%, 19%, 15%, and 6.5% of patients, respectively. Overall, 20% of patients had the early T-cell precursor (ETP)-LBL immunophenotype, as proposed by the WHO 2017 classification. For the early/pro-T/CD2(−) subtype, the five-year OS and EFS were 13% and 13%, while for all the other, non-pro-T subtypes, they were 69% and 67%. By multivariate analysis, only CD2(−) status and age > 35 years emerged as strong, independent factors influencing OS and EFS, while the risk of CR failure was influenced by age only (>35 years). (4) Conclusions: ETP was non-significant for OS, unless an ultra-high-risk pro-T/CD2(−) subtype was concerned.

B7-H6 is a new potential biomarker and therapeutic target of T-lymphoblastic lymphoma.

BackgroundB7-H6 is a novel co-stimulatory protein exclusively expressed on a variety of cancer cells and associated with poor prognosis. T-cell lymphoblastic lymphoma (T-LBL) is a highly aggressive hematological malignancy whose treatment requires reliable prognostic biomarkers and therapeutic targets. However, the rare nature and delayed progression of T-LBL have limited its clinical management.MethodsThe expression of B7-H6 was analyzed by immunohistochemistry (IHC) in 65 T-LBL samples; the association with the clinicopathological characteristics and prognosis was also investigated. B7-H6-depleted Jurkat cells were also generated to investigate the effect of B7-H6 on cell proliferation, migration, and invasion. RNA sequencing was used to explore differentially expressed genes.ResultsB7-H6 was expressed in 61.5% (40/65) of T-LBL patients; of note, 38.5% (25/65) of patients showed membrane/cytoplasmic expression of B7-H6. Although the expression of B7-H6 varied across samples and did not correlate with patient survival, it was significantly associated with B symptoms, high ECOG scores (3 to 4), elevated serum lactate dehydrogenase level, and reduced complete remission at interim evaluation. B7-H6 underwent translocation into the nucleus of T-LBL cells, showing a specific nuclear localization sequence in the C-terminus. Moreover, the depletion of B7-H6 in Jurkat cells impaired cell proliferation, migration, and invasion. RNAseq showed the differential expression of RAG-1, which may be involved in the tumorigenesis of T-LBL.ConclusionsB7-H6 may serve as a novel prognostic biomarker and therapeutic target of T-LBL.

Baseline Total Metabolic Tumor Volume and Total Lesion Glycolysis Measured on 18F-FDG PET-CT Predict Outcomes in T-Cell Lymphoblastic Lymphoma.

PurposeThere is no optimal prognostic model for T-cell lymphoblastic lymphoma (T-LBL). Here, we discussed the predictive value of total metabolic tumor volume (TMTV) and total lesion glycolysis (TLG) measured on 18F-fluorodeoxyglucose positron emission tomography–computed tomography (PET-CT) in T-LBL.Materials and MethodsThirty-seven treatment naïve T-LBL patients with PET-CT scans were enrolled. TMTV was obtained using the 41% maximum standardized uptake value (SUVmax) threshold method, and TLG was measured as metabolic tumor volume multiplied by the mean SUV. Progression-free survival (PFS) and overall survival (OS) were analyzed by Kaplan-Meier curves and compared by the log-rank test.ResultsThe optimal cutoff values for SUVmax, TMTV, and TLG were 12.7, 302 cm3, and 890, respectively. A high SUVmax, TMTV, and TLG indicated a shorten PFS and OS. On multivariable analysis, TMTV ≥ 302 cm3, and central nervous system (CNS) involvement predicted inferior PFS, while high SUVmax, TLG and CNS involvement were associated with worse OS. Subsequently, we generated a risk model comprising high SUVmax, TMTV or TLG and CNS involvement, which stratified the population into three risk groups, which had significantly different median PFS of not reached, 14 months, and 7 months for low-risk group, mediate-risk group, and high-risk group, respectively (p < 0.001). Median OS were not reached, 27 months, and 13 months, respectively (p < 0.001).ConclusionBaseline SUVmax, TMTV, and TLG measured on PET-CT are strong predictors of worse outcome in T-LBL. A risk model integrating these three parameters with CNS involvement identifies patients at high risk of disease progression.

The Expression of Paks and Its Clinical Significance in T-Cell Lymphoblastic Lymphoma

Background: T-cell lymphoblastic lymphoma (T-LBL) is a highly aggressive non-Hodgkin's lymphoma with poor prognosis and lacks of standard treatment approaches. PAK2, a member of the p21 activated kinase (PAKs) family, is a component of the gene-expression-based classifier which made great contributions to prognostic prediction of T-LBL(Cai et al. Leukemia 2020). This study aims to analyze the expression of PAKs in human T-LBL cell lines and tissues to clarify its clinical significance and evaluate the therapeutic activity of PAKs inhibitor in T-LBL. Methods: The mRNA and protein expression levels of PAKs in human T-lymphoid cell lines (H9) and T-LBL cell lines (Jurkat, SUP-T1 and CCRF-CEM) were detected by quantitative real-time PCR and western blot, respectively. We retrospectively collected 69 Formalin-fixed, Paraffin-embedded (FFPE) samples and corresponding clinical information from T -LBL patients between September 2000 and May 2015 at Sun Yat-sen University Cancer Center (SYSUCC). Affymetrix Human Gene 2.0 ST microarray (Thermo Fisher Scientific, Waltham, MA, USA) was used to detect the expression of PAKs. Mann-Whitney U test was used for comparing the expression differences between relapsed and non-relapsed patients. Cumulative relapse-free survival (RFS) time was calculated using the Kaplan-Meier method (Expression level higher than the upper quartile (P75) was defined as PAK1 or PAK 2 high expression. High expression (median as cutoff point) of both PAK1 and PAK2 was defined as PAK1/2 high expression). Pearson's chi-square test and Fisher's exact test were used to compare the distribution of clinical variables. Correlations between PAK1, PAK2 and NOTCH1 were evaluated using Spearman's correlation coefficient. Two PAK inhibitors, PF3758309 (PF) and FRAX597, were used to block PAK kinase activity pharmacologically. Cell viability was determined using the viability assay kit CCK-8. Cell cycle and cell apoptosis were analyzed by flow cytometry. All statistical analyses were performed using SPSS 24.0 software (SPSS, Armonk, NY, USA) or GraphPad Prism 8.0 (GraphPad, La Jolla, CA, USA). A P value &amp;lt; 0.05 was considered significant. The study protocol was approved by the Institutional Review Board of SYSUCC. Results: The mRNA and protein expression levels of PAK1 in T-LBL cell lines of Jurkat, SUP-T1 and CCRF-CEM cell lines were significantly higher than those in T-lymphoid cell lines H9 cell line (P&amp;lt;0.05) (Figure 1). Of the 69 T-LBL patients, 44 (63.8%) were male and the median age at diagnosis was 30 years (range: 16-44). The majority (72.5%) of the patients received acute lymphoblastic leukemia (ALL)-type chemotherapy regimens. The PAK2 mRNA expression level of 32 patients with relapsed disease was significantly higher than that of 37 cases without relapse (P=0.012), and no difference was found in mRNA expression of PAK1, 3, 4, 7(Figure 2). Patients with high PAK1 and PAK2 expression had significantly shorter median RFS than those with low PAK1 and PAK2 expression (PAK1 mRFS 31.5 months vs not reached(NR), HR=3.001, P=0.028; PAK2 mRFS 25.7 months vs NR, HR=3.981, P=0.027)(Figure 3). Patients with high PAK1/2 mRNA expression experienced earlier relapse than patients with low PAK1/2 mRNA expression (mRFS 26.0 months vs NR, HR=2.721, P=0.032)(Figure 3). PAK1/2 mRNA expression was found to be associated with hemoglobin concentration, Ki-67 expression, pleural and pericardia effusion, bone marrow involvement and chemotherapy response (P&amp;lt;0.05)(Table 1). Further, the PAK1 mRNA was correlated with the expression of NOTCH1, which is frequently mutated in T-LBL (r=0.5716, P&amp;lt;0.0001)(Figure 4). The PAK inhibitors PF and FRAX597 demonstrated strong anti-tumor activity in vitro (Table 2). Both inhibitors suppressed cell proliferation in a time- and dose-dependent manner (Figure 5). Both inhibitors induced cell cycle arrest in the G1/0 phase, accompanied by corresponding S phase reductions in all tested cells(Figure 6). The synergistic effect between PAK inhibitor PF and doxorubicin was also observed (Figure 7). Besides, PF could inhibit the phosphorylation of PAK1/2, Cyclin D1 and NF-κB in Jurkat cell line(Figure 8). Conclusions: PAK1 and PAK2 play certain roles in the occurrence and recurrence of T-LBL, and their potential as novel biomarkers deserves further exploring. Our results underscore the potential of PAK inhibitor as effective target therapy for T-LBL. Disclosures No relevant conflicts of interest to declare.

Event-free survival at 24months is a robust surrogate endpoint for long-term survival in pediatric, adolescent, and adult T cell lymphoblastic lymphoma.

T cell lymphoblastic lymphoma (T-LBL) has an aggressive clinical behavior. To date, powerful and consistent prognostic factors have not been established for T-LBL. In this study, we first evaluated the association of event-free survival (EFS) at 24months (EFS24) with overall survival (OS) in T-LBL patients. Besides, we sought to identify clinical factors of prognostic importance in this rare entity. Between January 2006 and December 2017, ninety-one patients with newly diagnosed T-LBL were retrospectively analyzed. EFS was defined as the time from diagnosis to relapse or progression, unplanned retreatment, death from any cause, or to the last follow-up. In total, 91 patients with a median age of 24years were enrolled. At a median follow-up of 40.4months (range, 1.4 to 163.3months), the 5-year OS and EFS was 47.9% and 43.2%, respectively. Of all patients, 45 (49.5%) achieved EFS24 and 46 (50.5%) did not. Patients who achieved EFS24 showed a markedly superior outcome, compared with those who failed to achieve EFS24 (5-year OS, 90.5% vs 3%, P< 0.001). Univariate analysis indicated bone marrowinvolvement, response to induction treatment, and stem cell transplantation (SCT) consolidation to be prognostic factors for EFS and OS. In addition, compared with the patients receiving non-Hodgkin's lymphoma (NHL)-like treatmentprotocols, patients treated with hyper-CVAD showed significantly improved EFS and OS. Such survival advantage in terms of EFS and OS was also observed of BMF-90 regimens over NHL-like therapy, despite that the difference in EFS did not reach statistical significance (P= 0.056). Multivariate analysis demonstrated that achievement of complete remission (CR) after induction therapy and SCT consolidation were independent prognostic indicators for both EFS and OS. We confirm that EFS24 is a strong surrogate endpoint for long-term survival in T-LBL, which is clinically useful for individualized risk reassessment, future clinical trial design, and biomarker discovery validation. Further validation in the context of directed prospective clinical trials is warranted.

BRD2 induces drug resistance through activation of the RasGRP1/Ras/ERK signaling pathway in adult T-cell lymphoblastic lymphoma.

BackgroundAdult patients with T‐cell lymphoblastic lymphoma (T‐LBL) are treated with high‐intensity chemotherapy regimens, but the response rate is still unsatisfactory because of frequent drug resistance. We aimed to investigate the potential mechanisms of drug resistance in adults with T‐LBL.MethodsGene expression microarray was used to identify differential mRNA expression profiles between chemotherapy‐resistant and chemotherapy‐sensitive adult T‐LBL tissues. Real‐time PCR and immunohistochemistry were performed to detect the expression of bromodomain‐containing protein 2 (BRD2) and c‐Myc in fresh‐frozen T‐LBL tissues from 85 adult patients. The Ras pull‐down assay was performed to monitor Ras activation. Chromatin immunoprecipitation assays were used to analyze the binding of E2F transcription factor 1 (E2F1)/BRD2 to the RAS guanyl releasing protein 1 (RasGRP1) promoter region. The drug resistance effect and mechanism of BRD2 were determined by both in vivo and in vitro studies.ResultsA total of 86 chemotherapy resistance‐related genes in adult T‐LBL were identified by gene expression microarray. Among them, BRD2 was upregulated in chemotherapy‐resistant adult T‐LBL tissues and associated with worse progression‐free survival and overall survival of 85 adult T‐LBL patients. Furthermore, BRD2 suppressed doxorubicin (Dox)‐induced cell apoptosis both in vitro and in vivo. The activation of RasGRP1/Ras/ERK signaling might contribute to the Dox resistance effect of BRD2. Besides, OTX015, a bromodomain and extra‐terminal (BET) inhibitor, reversed the Dox resistance effect of BRD2. Patient‐derived tumor xenograft demonstrated that the sequential use of OTX015 after Dox showed superior therapeutic effects.ConclusionsOur data showed that BRD2 promotes drug resistance in adult T‐LBL through the RasGRP1/Ras/ERK signaling pathway. Targeting BRD2 may be a novel strategy to improve the therapeutic efficacy and prolong survival of adults with T‐LBL.