T-acute Lymphoblastic Leukemia Research Articles

Pharmacodynamic and Toxicity Studies of 6-Isopropyldithio-2'-guanosine Analogs in Acute T-Lymphoblastic Leukemia.

(1) Background: The research group has developed a new small molecule, 6-Isopropyldithio-2'-deoxyguanosine analogs-YLS004, which has been shown to be the most sensitive in acute T-lymphoblastic leukemia cells. Moreover, it was found that the structure of Nelarabine, a drug used to treat acute T-lymphoblastic leukemia, is highly similar to that of YLS004. Consequently, the structure of YLS004 was altered to produce a new small molecule inhibitor for this study, named YLS010. (2) Results: YLS010 has exhibited potent anti-tumor effects by inducing cell apoptosis and ferroptosis. A dose gradient was designed for in vivo experiments based on tentative estimates of the toxicity dose using acute toxicity in mice and long-term toxicity in rats. The study found that YLS010 at a dose of 8 mg/kg prolonged the survival of late-stage acute T-lymphoblastic leukemia mice in the mouse model study. (3) Conclusions: YLS010 has demonstrated specific killing effects against acute T-lymphoblastic leukemia both in vivo and in vitro. Preclinical studies of YLS010 offer a new opportunity for the treatment of patients with acute T-lymphoblastic leukemia in clinical settings.

Investigates the genetic types of acute lymphoblastic leukemia in full remission Indonesian patients: a cross-sectional analysis

Acute lymphoblastic leukemia (ALL) is most common in children, especially those aged 1–4 years old, and the second most common acute leukemia in adults. The updated International Consensus Classification (ICC) of B-acute lymphoblastic leukemia (B-ALL) and T-acute lymphoblastic leukemia (T-ALL) includes recent clinical, cytogenetic, and molecular data. Transcriptome sequencing (RNA-seq) was performed on 200 bone marrow specimens using TruSeq library preparation and HiSeq 2000/2500 or NovaSeq 6000 sequencers (Illumina). The purpose of this study was to define the frequency of chromosomal abnormalities of ALL patients in adults and children in Indonesian patients after full remission for international collaboration has improved and advanced the diagnosis and treatment of ALL in Indonesia. Our resulting data showed that the most common structural abnormality was t(9;22) in 15% of the patients. The frequency of genetic abnormalities was 69 % and 60% for numerical and/or structural in the B-ALL and T-ALL patients, respectively. The adults had a higher incidence of t(9;22) and a lower incidence of hyperdiploid than children. In conclusion: The results of this study molecular subtypes differ strikingly in their responses to treatment that needs more assessment.

Exploring novel protein biomarkers for early-stage diagnosis and prognosis of T-acute lymphoblastic leukemia (T-ALL)

BackgroundEfficient classification of T-acute lymphoblastic leukemia (T-ALL) involves considering various factors, such as age, white blood cell count, and chromosomal alterations. However, studying protein markers are crucial to improving T-ALL patients' diagnosis and treatment. A study analyzing the expression of proteomes was conducted to identify promising early-stage biomarkers for T-ALL patients MethodsLabel-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to analyze the blood proteins of both patients and healthy individuals to identify new biomarkers for T-ALL. The findings were validated by RT-PCR, ELISA and computational analysis ResultsThe study identified 1467 proteins in the blood, of which nine were upregulated and 35 were downregulated by more than 2-fold. T-ALL patients showed a significant increase in specific disease-related proteins, such as eleven-nineteen lysine-rich leukemia protein, triggering receptor expressed on myeloid cells 1, cisplatin resistance-associated-overexpressed protein, X-ray radiation resistance-associated protein 1, tumor necrosis factor receptor superfamily member 10D, protein S100-A8, and copine-4, by more than 3-fold ConclusionThe findings of this study provide a valuable protein map of leukemic cells and identify potential biomarkers for leukemic aggressiveness. However, further studies using larger T-ALL patient samples must confirm these preliminary results.

[Resveratrol Inhibits T-acute Lymphoblastic Leukemia in Mice by Regulating Notch1 Signaling Pathway].

To observe the effect of resveratrol (Res) on T-acute lymphoblastic leukemia (T-ALL) mice, and further explore its mechanism on Notch1 signaling pathway. Twenty-five 6-8 weeks old female C57BL/6 mice were randomly divided into control group, T-ALL group and Res group. Res group was further divided into low-Res, middle-Res and high-Res group. The percentage of leukemia cells in peripheral blood and spleen cell suspension were detected by flow cytometry and Wright-Giemsa staining, pathological morphology of spleen and bone marrow tissues were observed by HE staining, the expression levels of Notch1, Hes-1, c-Myc, miR-19b and PTEN mRNA in spleen tissue were detected by RT-qPCR, and the protein levels of Notch1, Hes-1, c-Myc, p-PTEN and PTEN were detected by Western blot. Compared with control group, the leukemia cells in peripheral blood of mice in T-ALL group were markedly increased, accompanied by diffuse infiltration of leukemia cells in spleen and bone marrow tissues, the mRNA levels of Notch1, Hes-1, c-Myc, miR-19b and the protein levels of Notch1, Hes-1, c-Myc were increased (P <0.01), while the expression of PTEN mRNA and protein were significantly decreased in the spleen tissue of T-ALL mice (P <0.01). The above indicators in the H-Res group were reversed compared with T-ALL group after administration of resveratrol. Resveratrol may play a role in anti T-ALL by inhibiting Notch1 signaling pathway in mice.

An Anti-CD7 Antibody-Drug Conjugate Target Showing Potent Antitumor Activity for T-Lymphoblastic Leukemia (T-ALL).

Acute T-lymphoblastic leukemia (T-ALL) is a type of leukemia that can occur in both pediatric and adult populations. Compared to acute B-cell lymphoblastic leukemia (B-ALL), patients with T-cell T-ALL have a poorer therapeutic efficacy. In this study, a novel anti-CD7 antibody-drug conjugate (ADC, J87-Dxd) was successfully generated and used for T-ALL treatment. Firstly, to obtain anti-CD7 mAbs, we expressed and purified the CD7 protein extracellular domain. Utilizing hybridoma technology, we obtained three anti-CD7 mAbs (J87, G73 and A15) with a high affinity for CD7. Both the results of immunofluorescence and Biacore assay indicated that J87 (KD = 1.54 × 10-10 M) had the highest affinity among the three anti-CD7 mAbs. In addition, an internalization assay showed the internalization level of J87 to be higher than that of the other two mAbs. Next, we successfully generated the anti-CD7 ADC (J87-Dxd) by conjugating DXd to J87 via a cleavable maleimide-GGFG peptide linker. J87-Dxd also possessed the ability to recognize and bind CD7. Using J87-Dxd to treat T-ALL cells (Jurkat and CCRF-CEM), we observed that J87-Dxd bound to CD7 was internalized into T-ALL cells. Moreover, J87-Dxd treatment significantly induced the apoptosis of Jurkat and CCRF-CEM cells. The IC50 (half-maximal inhibitory concentration) value of J87-Dxd against CCRF-CEM obtained by CCK-8 assay was 6.3 nM. Finally, to assess the antitumor efficacy of a J87-Dxd in vivo, we established T-ALL mouse models and treated mice with J87-Dxd or J87. The results showed that on day 24 after tumor inoculation, all mice treated with J87 or PBS died, whereas the survival rate of mice treated with J87-Dxd was 80%. H&E staining showed no significant organic changes in the heart, liver, spleen, lungs and kidneys of all mice. In summary, we demonstrated that the novel anti-CD7 ADC (J87-Dxd) had a potent and selective effect against CD7-expressing T-All cells both in vitro and in vivo, and could thus be expected to be further developed as a new drug for the treatment of T-ALL or other CD7-expression tumors.

Post-Allograft Romidepsin Maintenance Mitigates Relapse Risk and Stimulates the Graft-Versus-Malignancy Effect through Enhanced NK-Cell Cytotoxicity in Patients with T-Cell Malignancies: Final Results of a Phase I/II Trial

Background: For patients with high risk, and relapsed/refractory T-cell malignancies allogeneic stem cell transplant (allo-SCT) is the only available potentially curative therapy. The efficacy of allo-SCT is limited in this population by high rates of relapse, with rates up to 55-60% post-allo-SCT. We designed a phase I/II trial, evaluating the combination of the histone deacetylases inhibitor romidepsin (rom) with busulfan/fludarabine (BuFlu) conditioning, followed by romidepsin maintenance (m-rom) in patients receiving allo-SCT for T-cell malignancies (NCT02512497). Here we present final clinical results of this therapeutic approach, including an evaluation of the stimulatory effects of m-rom on the graft-versus-malignancy effect through NK-cells post-allo-SCT. Methods: This was a phase I/II clinical trial. Eligible patients had: a diagnosis of T-cell leukemia (including T-acute lymphoblastic leukemia) or T-cell lymphoma (TCL, cutaneous or peripheral) in at least a partial remission requiring an allo-SCT, &amp;lt;70 years of age, with a matched sibling/unrelated donor. Patients received myeloablative (20K) or reduced intensity (16K) BuFlu with rom, followed by standard tacrolimus/methotrexate GVHD prophylaxis and anti-thymocyte globulin for unrelated donors (MUD). An expansion cohort of up to 30 patients (total) was included. M-rom was initiated between day +28 and +100 for 1 year (2 nd year optional), with built in dose reductions for toxicity. An efficacy endpoint of 20% reduction in relapse risk at 1-year in the whole cohort or any individual disease cohort was set based upon historical/CIBMTR (Center for Blood and Marrow Transplant Research) controls was pre-specified. The effect of m-rom on NK-cell cytotoxicity was assessed on samples taken pre-transplant, and 1, 3, 6, 12 months post allo-SCT. NK cytotoxicity was assessed by isolating mononuclear cells from patient samples at each timepoint, comparing those on m-rom (n=13) versus those who did BuFlu controls who did not receive m-rom (n=16). Cells were targeted against K562 targets using the calcein-AM assay. K-M and Fine-Gray models were used to estimate PFS, OS, and cumulative incidence, and compare survival across groups. Results: 28 patients were enrolled on this trial (Table). With a median follow-up time of 15 months, the median OS is 3.3 years (95% CI: 0.85-not reached), with a 1 and 3 year OS probability of 68% and 54%. The median PFS is 2.3 years (95% CI: 0.6-not reached), with 1 and 3-year PFS of 61% and 41%. Cumulative incidence (CI) of NRM at day 100 and 1 year were 14.3% and 21.4%. CI of grade II-IV aGHVD and extensive cGVHD were 46.4% and 38%. The CI of relapse (CIR) was 18.1% at 1 year and 33% at 3 -years. There was no difference between PFS among patients with MRD versus those without MRD prior to transplant (p=0.43). The CIR of patients with TCL was 9.3% at 1 and 3 years, whereas the CIR of patients with leukemia was 25% at 1 year and 48% at 3 years. No patients with PTCL relapsed, 1/2 patients with CTCL relapsed, and 4/6 patients with T-PLL are alive, disease free. Pre-specified general CIR was 55% for all patients. With an overall CIR of 18% at 1 year, and 33% at 3-years, this trial met its pre-specified endpoint of decreasing relapse by 20% at 1-year, and performed well compared to published CIBMTR control populations. Matched propensity score analysis across disease subsets with CIBMTR controls will be presented. 18/28 (64%) of patients received m-rom with a median number of 13 cycles (range 1-47). 8 patients experienced grade 3/4 adverse events (AE), and 1 patient discontinued m-rom due to toxicity, unlikely related to romidepsin (loss of CD3 chimerism). When NK-cytotoxicity was assessed between the two groups after starting maintenance, NK-cytotoxicity in the m-rom group was significantly higher than in those without m-rom (p&amp;lt;0.0001) (Figure). Conclusions: BuFluRom with m-rom is effective at decreasing relapse in patients with T-cell malignancies, with 1-year CI relapse below expected relapse rates for this set of diseases, meeting the pre-specified efficacy endpoint. NK-cell cytotoxicity data in a large sample set of trial patients demonstrates that m-rom enhances NK-cell cytotoxicity post allo-SCT, augmenting the GVL effect and likely accounting for at a minimum, some of the decrease in relapse seen on this trial. M-rom should be considered a new option post allo-SCT to mitigate relapse in patients with T-cell malignancies.

Evaluation of aberrant expression of CD markers in acute leukemia cells

Background and objective: Worldwide immunophenotyping by flow cytometry (FCM) in acute leukemia (AL) is the golden step in the diagnosis. It’s very common for acute leukemias to aberrantly express antigens or cluster of differentiation (CD) markers which are usually expressed in other lineages of the disease hence this study aimed at determining the prevalence of aberrancy in AL and to find out the frequency of each aberrant CD marker and their association with the clinic-hematological profile of the cases. Methods: Following history and clinical examination of enrolled patients, blood and/or bone marrow aspirate was drawn for morphological examination and immunophenotyping by FCM from 86 newly diagnosed acute leukemia cases then multiple steps procedure was applied followed by interpretation of the results. Results: The prevalence of aberrant phenotype was 46.5%. The proportional frequency of aberrant phenotype in acute myeloid leukemia (AML) was 41%, in B-acute lymphoblastic leukemia (B-ALL) was 48.8% and in T-acute lymphoblastic leukemia (T-ALL) was 66.6%. The commonest aberrant CD markers in AML were CD22 and CD2, in B-ALL were CD66c and CD13 while in T-ALL were CD13 and CD33. The aberrant phenotype harbored lower white blood cell (WBC) count and blast percentage in PB, also splenomegaly was more frequent in lymphoid positive (Ly+) AML and myeloid positive (My+) T-ALL while in B-ALL, splenomegaly was more frequent in myeloid negative (My-) B-ALL. Conclusion: Aberrant phenotype prevalence in our study sample was comparable to other studies, considerable frequency of aberrant markers is present in cases of AL and some variations exist regarding the clinical and hematological profile of the aberrant group.

Dual Targeting of Apoptotic and Signaling Pathways in T-Lineage Acute Lymphoblastic Leukemia.

Relapsed T-acute lymphoblastic leukemia (T-ALL) has limited treatment options. We investigated mechanisms of resistance to BH3 mimetics in T-ALL to develop rational combination strategies. We also looked at the preclinical efficacy of NWP-0476, a novel BCL-2/BCL-xL inhibitor, as single agent and combination therapy in T-ALL. We used BH3 profiling as a predictive tool for BH3 mimetic response in T-ALL. Using isogenic control, venetoclax-resistant (ven-R) and NWP-0476-resistant (NWP-R) cells, phosphokinase array was performed to identify differentially regulated signaling pathways. Typical T-ALL cells had increased dependence on BCL-xL, whereas early T-precursor (ETP)-ALL cells had higher BCL-2 dependence for survival. BCL-2/BCL-xL dual inhibitors were effective against both subtypes of T-lineage ALL. A 71-protein human phosphokinase array showed increased LCK activity in ven-R cells, and increased ACK1 activity in ven-R and NWP-R cells. We hypothesized that pre-TCR and ACK1 signaling pathways are drivers of resistance to BCL-2 and BCL-xL inhibition, respectively. First, we silenced LCK gene in T-ALL cell lines, which resulted in increased sensitivity to BCL-2 inhibition. Mechanistically, LCK activated NF-κB pathway and the expression of BCL-xL. Silencing ACK1 gene resulted in increased sensitivity to both BCL-2 and BCL-xL inhibitors. ACK1 signaling upregulated AKT pathway, which inhibited the pro-apoptotic function of BAD. In a T-ALL patient-derived xenograft model, combination of NWP-0476 and dasatinib demonstrated synergy without major organ toxicity. LCK and ACK1 signaling pathways are critical regulators of BH3 mimetic resistance in T-ALL. Combination of BH3 mimetics with tyrosine kinase inhibitors might be effective against relapsed T-ALL.

Clinical features, laboratory characteristics, and outcome of ETP and TCRA/D aberrations in pediatric patients with T-acute lymphoblastic leukemia

BackgroundT-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy with few accepted prognostic factors that limit the efficiency of therapy. The aim of the current study was to assess the clinical and laboratory features of T-cell receptor (TCR) aberrations and early T-cell precursor (ETP) subtype as well as their outcome to therapy.MethodsSixty-three newly diagnosed pediatric T-ALL patients were assessed for the ETP status using immunophenotyping. Screening of TCRA/D aberrations was done by fluorescent in situ hybridization (FISH). The data were correlated to the patients’ clinical features, response to treatment, and survival rates.ResultsSeven patients (11%) had ETP-ALL. The ETP-ALL patients were older (P = 0.013), presented with lower white blood cell (WBC) count (P = 0.001) and lower percentage of peripheral blood (PB) blast cells (P = 0.037), more likely to have hyperdiploid karyotype (P = 0.009), and had been associated with TCRA/D gene amplification (P = 0.014) compared to other T-ALL patients. Of note, the same associations had been significantly observed in patients with TCRA/D gene amplification. Patients with TCRA/D amplification frequently coincided with TCRβ aberrations (P = 0.025). TCR-β aberrations were significantly associated with negative MRD at the end of induction compared to TCR-β-negative patients. There was a nonsignificant trend of ETP-positive cases to have lower overall survival (OS) (P = 0.06). Patients with TCR aberrations had no significant differences regarding disease-free survival (DFS) or OS rates compared to those with normal TCR.ConclusionETP-ALL patients tend to have increased mortalities. There was no significant impact of TCR aberrations on the survival rates of the patients.

Treatment outcomes for acute T-lymphoblastic leukemias/lymphomas: data from the ALL-2016 multicenter prospective randomized trial

Background . The effectiveness of therapy for acute T-lymphoblastic leukemia (T-ALL) and T-lymphoblastic lymphoma (T-LBL) has significantly improved over the past decades, including through the implementation of hematopoietic stem cell transplantation (HSCT). The Russian multicenter ALL-2009 study (ClinicalTrials.gov NCT01193933) showed that performing autologous HSCT (auto-HSCT) in patients with T-ALL/LBL improves long-term results. However, the study was non-randomized and the need for auto-HSCT in clinical practice requires careful study. Aim . To assess the significance of auto-HSCT in patients with T-ALL/LBL in the framework of ALL-2016 multicenter, prospective, randomized study (ClinicalTrials.gov NCT03462095). Materials and methods . The study included 109 adult patients with T-ALL/LBL (m:f 82:27). Median age was 31 (18– 52) years. T-ALL was diagnosed in 88 (81 %) patients, T-LBL in 21 (19 %) patients. All patients are treated according to the ALL-2016 protocol. Using the web platform, upon completion of induction therapy (+70 days), all T-ALL/LBL patients who achieved clinical and hematological remission were randomized to the auto-HSCT arm or chemotherapy alone (CT). Centralized monitoring of minimal residual disease (MRD) in bone marrow samples was performed by multicolor flow cytometry at the control points according to the ALL-2016 protocol. Statistical analysis was performed using SAS 9.4. Results . 87 patients with T-ALL/LBL were randomized: 44 to the auto-HSCT arm and 43 to the CT arm. Further analysis included 25 patients who underwent auto-HSCT and 36 patients receiving only CT. Three-year relapse-free survival in T-ALL is estimated at 62 % (auto-HSCT) vs. 81 % (CT) (p = 0.3422), and in T-LBL – 67 % (auto-HSCT) vs. 79 % (CT) (p = 0.59). MRD persistence on the +70 th day of therapy according to the ALL-2016 protocol was determined in 40 % of patients (autoHSCT) and in 67 % (CT) (p = 0.057). In a multivariate analysis, it was determined that T-ALL from early T-cell precursor (ETP-variant) and MRD persistence after the end of II phase induction are the main risk factors for relapse in the treatment according to the ALL-2016 protocol. Conclusion . Performing auto-HSCT both in patients with T-ALL and T-LBL, and with MRD persistence on day +70 according to the ALL-2016 protocol did not improve long-term results. The development of new programs for the treatment of patients with MRD persistence, as well as the ETP variant of T-ALL, is of current interest.

The pathogenesis and development of targeted drugs in acute T lymphoblastic leukaemia.

Acute lymphoblastic leukaemia (ALL) is mainly classified into acute T- and B-lymphoblastic leukaemia according to the source of its lymphocytes, thymus and bone. Among them, the incidence of adult T-cell accounts for about 25% of adult acute lymphoblastic leukaemia, but the degree of malignancy is high and the treatment rate and prognosis are poor. At this stage, there are few targeted drugs and the commonly used broad-spectrum chemotherapeutic drugs have poor efficacy and many adverse drug reactions. Understanding and investigating the pathogenesis of T-acute lymphoblastic leukaemia is very important for further developing new targeting drugs and improving existing drugs. Dysregulated signalling pathways are the main aetiological factors of T-acute lymphoblastic leukaemia. They play crucial roles in promoting tumour initiation, progression, drug design and therapy responses. This is primarily because signalling pathways are indispensable for many cellular biological processes, including tumour growth, migration, invasion, metastasis and others. As a result, small molecule inhibitors targeting the major kinase components of the signalling pathway have received a lot of attention and have been developed and evaluated in preclinical models and clinical trials. Already marketed drugs are also being repurposed in combination therapies to further improve efficacy and overcome tumour cell resistance. In this review, we have aimed to examine the latest and most classical signalling pathways in the aetiology of T-acute lymphoblastic leukaemia and shed light on potential targets for novel therapeutic agents to act on.

IL-15 Prevents the Development of T-ALL from Aberrant Thymocytes with Impaired DNA Repair Functions and Increased NOTCH1 Activation.

We previously reported that NOD.Scid mice lacking interleukin-15 (IL-15), or IL-15 receptor alpha-chain, develop T-acute lymphoblastic leukemia (T-ALL). To understand the mechanisms by which IL-15 signaling controls T-ALL development, we studied the thymocyte developmental events in IL-15-deficient Scid mice from NOD and C57BL/6 genetic backgrounds. Both kinds of mice develop T-ALL characterized by circulating TCR-negative cells expressing CD4, CD8 or both. Analyses of thymocytes in NOD.Scid.Il15-/- mice prior to T-ALL development revealed discernible changes within the CD4-CD8- double-negative (DN) thymocyte developmental stages and increased frequencies of CD4+CD8+ double-positive cells with a high proportion of TCR-negative CD4+ and CD8+ cells. The DN cells also showed elevated expressions of CXCR4 and CD117, molecules implicated in the expansion of DN thymocytes. T-ALL cell lines and primary leukemic cells from IL-15-deficient NOD.Scid and C57BL/6.Scid mice displayed increased NOTCH1 activation that was inhibited by NOTCH1 inhibitors and blockers of the PI3K/AKT pathway. Primary leukemic cells from NOD.Scid.Il15-/- mice survived and expanded when cultured with MS5 thymic stromal cells expressing Delta-like ligand 4 and supplemented with IL-7 and FLT3 ligand. These findings suggest that IL-15 signaling in the thymus controls T-ALL development from aberrant thymocytes with an impaired DNA repair capacity and increased NOTCH1 activation.

A Comparative Analysis of Early T-Cell Precursor Lymphoblastic Leukemia/Lymphoma (ETP-ALL/LBL) and Non-ETP-ALL/LBL in a Tertiary Cancer Care Center Based in Western India.

Early T-cell precursor lymphoblastic leukemia (ETP-ALL) has a unique immunophenotype with very early T-cell differentiation. The current study summarises the distinct clinicopathological aspects of ETP-ALL and compares them with non-ETP-ALL. Twenty-nine ETP-ALL and 191 non-ETP-ALL cases were retrieved between 2018 and 2021. A P value was determined for each of the patient charaterisics (Table 1) to see for any significant relationship (P < 0.05) with ETP-ALL versus non-ETP-ALL. Kaplan-Meier log rank test was applied to look for any significant differences in OS for both the ALLs. ETP-ALL had an incidence of 12.6% out of total T-Acute lymphoblastic leukemia (T-ALL/LBL) in the past 3-years. Compared to non-ETP-ALL, ETP-ALL cases were associated with lower median age and male-to-female ratio. There was no statistically significant difference in the complete remission rate between both the subtypes. ETP-ALL was seen to be associated with high induction failure and relapse rate compared to non-ETP-ALL. To summarise, since the 2-year OS was poor compared to western research (for both ALLs), an intensive chemo-regimen should be implemented in the current situation. Some unusual markers were observed on flow-cytometry (ETP-ALL), which can be useful for MRD quantification, prognosis, and further trials for newer targeted therapies.

Aleukemic Leukemia: A Rare Presentation

Acute leukemia can present as leukemic blast in peripheral blood and bone marrow. Aleukemic leukemia is a form of leukemia that occurs without detectable levels of abnormal leukocytes within the blood. We put forth a unique case of aleukemic leukemia presenting with persistent fever that was diagnosed using immunophenotyping and flow cytometry as acute T-Lymphoblastic Leukemia.

Leukemia-associated aberrant immunophenotype: A flow cytometry-based experience of 110 cases from a tertiary care center in Northern India.

Leukemic cells express a characteristic set of "cluster of differentiation" (CD) markers, which forms the basis of the current WHO classification. Leukemia-associated aberrant immunophenotype (LAIP) refers to expression of unusual CD markers by leukemic cells, which are not normally expressed by their respective lineage. The incidence of LAIP varies considerably, and its clinical implications, prognostic relevance, and sensitivity to therapy are still debatable. This study was conducted to identify the immunophenotypic aberrancies in newly diagnosed leukemias in our Institute. This was an observational study, which included newly diagnosed leukemias on flow cytometry. Aberrant immunophenotypic expressions were recorded whenever present and were correlated with prognostic factors like age, gender, and total leucocyte count (TLC). The study included 110 newly diagnosed cases of leukemias (85 acute and 25 chronic) over 1.5 years. Immunophenotypic aberrancies were detected in 40.4% of the cases. The highest incidence of aberrations was detected in acute myeloid leukemia (60.7%). LAIPs were detected in 50% of T-acute lymphoblastic leukemia and 25% cases of in B-cell acute lymphoblastic leukemia (B-ALL). Aberrant CD33 and CD56 expression in B-ALL correlated with poor prognostic factors like higher age and higher TLC, respectively. Immunophenotypic aberrancies were present in 28% cases of chronic lymphocytic leukemia. The results of this study have generated valuable baseline data on the incidence of LAIPs in this region. This information is vital because establishing LAIPs at the time of diagnosis is crucial for disease monitoring. Some LAIPs are associated with underlying cytogenetic abnormalities and hence impact the management and prognosis.

International Consensus Classification of acute lymphoblastic leukemia/lymphoma.

The updated International Consensus Classification (ICC) of B-acute lymphoblastic leukemia (B-ALL) and T-acute lymphoblastic leukemia (T-ALL) includes both revisions to subtypes previously outlined in the 2016 WHO classification and several newly described entities. The ICC classification incorporates recent clinical, cytogenetic, and molecular data, with a particular emphasis on whole transcriptome analysis and gene expression (GEX) clustering studies. B-ALL classification is modified to further subclassify BCR::ABL1-positive B-ALL and hypodiploid B-ALL. Additionally, nine new categories of B-ALL are defined, including seven that contain distinguishing gene rearrangements, as well as two new categories that are characterized by a specific single gene mutation. Four provisional entities are also included in the updated B-ALL classification, although definitive identification of these subtypes requires GEX studies. T-ALL classification is also updated to incorporate BCL11B-activating rearrangements into early T-precursor (ETP) ALL taxonomy. Additionally, eight new provisional entities are added to the T-ALL subclassification. The clinical implications of the new entities are discussed, as are practical approaches to the use of different technologies in diagnosis. The enhanced specificity of the new classification will allow for improved risk stratification and optimized treatment plans for patients with ALL.

Correlation of Minimal Residual Disease (MRD) at the End of Induction (EOI) and Event Free Survival (EFS) in T-Cell Lymphoblastic Lymphoma (T-LL), a Report from the Children's Oncology Group (COG) Trial AALL1231

Background: Defining prognostic variables that correlate to EFS in T-LL remains a clinical challenge. Traditional variables such as stage or radiologic response to therapy have failed to correlate with EFS in recent trials. AALL1231 was a COG phase 3 clinical trial for newly diagnosed with T Acute Lymphoblastic leukemia (T-ALL) or T-LL patients that randomized children and young adults (age 1-30 years) to a modified augmented BFM (aBFM) backbone with no bortezomib (Arm A) or with bortezomib (Arm B) during induction and delayed intensification (DI) (1.3mg/m2 x 4 doses per block). T-LL patients were stratified as standard (SR), intermediate (IR), or very high risk (VHR), based upon minimal detectable disease (MDD) in the bone marrow at diagnosis, prior steroid pre-treatment and radiographic response at the end of induction. Bone marrow (BM) samples to assess MRD by flow cytometry at the EOI were an optional submission for T-LL participants and those collected were analyzed to assess its correlation to EFS. Results: AALL1231 accrued 209 T-LL patients from 2014 until closure in 2017. At the end of induction, 43.6% were in radiologic remission, 55.4% had a partial response and 1% had stable disease or no response. There were 86 (41%) patients for whom EOI samples for MRD assessment were submitted. Patients with MRD <0.1% (n=75) at EOI had a superior EFS versus those with MRD >0.1% (n= 11), (89.0 + 4.4% verses 63.6 + 17.2%, p= 0.025). Overall survival did not significantly differ between the two groups (88.9 + 4.4% versus 72.7 + 15.5% p= 0.15). As previously reported, Arm B demonstrated a superior outcome for both EFS and OS for the entire cohort. Within risk groups, IR and SR patients had similar EFS outcomes (Arm A 73.9 + 7.5% versus 80.4 + 6.7, Arm B 87.2 + 5.8 % versus 90.5 + 4.8%) with only two patients the VHR, both in Arm B. Cox regression for EFS using Arm A as a reference demonstrated that MRD EOI ≥0.1% was associated with a greater risk of inferior outcome (Hazard Ratio, HR = 3.73 (1.12-12.40, p = 0.032), (Table 1) which was independent of treatment arm assignment. OS failed to reach statistical significance for patients MRD EOI ≥0.1%, HR = 2.714 (0.72-10.44, p=0.14). Furthermore, Cox regression failed to demonstrate a superior EFS with decreasing MDD at diagnosis (>5% versus 1-5%, versus <1%), HR=2.67 (0.336-21.145, p=0.141, HR= 0.830 (0.255-2.699, HR= 0.57 (0.289-1.073, p= 0.080)(Table 2). Conclusions: In this pediatric T-LL trial, BM EOI MRD by flow cytometry was associated with improved EFS in in both arms (+/- bortezomib) in both univariate and multivariate analyses. In contrast, race, age, gender, risk group, MDD, stage and radiologic response to therapy were not prognostic. Consideration to incorporate MRD at EOI into the design of future trials will establish its value in defining risk groups for subsequent therapeutic trials. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Pediatric Acute Lymphoblastic Leukemia with Pdgfrb Fusions: A Multicenter Retrospective Study

Objective: PDGFRB fusions in acute lymphoblastic leukemia (ALL) is rare. PDGFRB fusions was mainly observed in B-ALL, but also in T-ALL. Previous studies were mostly limited to anecdotal cases, suggesting that this subtype of ALL had low rate of remission and poor prognosis. The addition of tyrosine kinase inhibitor (TKI) to intensive chemotherapy may improve the outcome, but lack of long-term survival data. Herein, we retrospectively analyzed the clinical characteristics, survival and prognostic factors for 28 pediatric patients with PDGFRB fusions ALL. Methods: This multicenter, retrospective study included pediatric patients with newly diagnosed PDGFRB fusion ALL after treatments according to the CCCG-ALL-2015 and CCCG-ALL-2020 protocols from April 2015 to April 2022 in 10 hospitals in China. All patients completed the followed up. The baseline characteristics and treatment outcomes of these patients, including complete remission, drug resistance, relapse, treatment-related mortality and minimal residual disease (MRD) at end of induction therapy, as well as hematopoietic stem cell transplantation (HSCT) were collected. The outcome was compared between patients treated with and without TKI. Fisher test and nonparametric rank sum test were used for comparative analysis. 4-year event-free survival (EFS) and 4-year overall survival (OS) were estimated by use of Kaplan-Meier methods, and the 4-year cumulative incidence of relapse was calculated by use of a competing risk model gray test. Univariate and multivariate analyses were performed by use of Cox regression. Results: We identified 28 pediatric patients with newly diagnosed PDGFRB fusion ALL, which accounted for approximately 0.5% of childhood ALL. In all 28 patients, 21 (75%) were male and 7 (25%) were female; the median age was 10 (1-16) years old; median white blood cell (WBC) was 29.8 (1.6-627.7) × 10^9/L; 24 (85.7%) were acute B-lymphoblastic leukemia and 4 (14.3%) were acute T-lymphoblastic leukemia; one case (3.5%) was observed accompanied by the CNS involvement at diagnosis. Twenty of the patients received TKI treatment. Only 6 patients underwent hematopoietic stem cell transplantation. For all 28 patients, the median follow-up was 25.5 (3-77) months, the 4-year EFS was 29.2% (95% CI, 13.6-36.4), the 4-year OS was 42.1% (95% CI, 30.6-61.4), and the 4-year cumulative incidence of relapse was 58% (95% CI, 45·6-70·1). Patients in TKI group had a high rate of complete remission after induction therapy compared with those in no-TKI group (90% vs 63.6%). However, no significant differences were observed in the 4-year EFS and 4-year OS between TKI group and no-TKI group (EFS, 21.1% vs 43.8%, P=0.925; OS, 50.1% vs 36.5%, P=0.807). The cumulative incidences of relapse between these two groups also have no remarkable differences (P >0.05). Among the 6 patients who underwent HSCT, 2 patients who achieved CR1 survived and 4 died. Univariate analysis showed that minimal residual disease (MRD) ≥10-4 after induction therapy for D46 and IKZF1 positive had a poor prognosis (all P <0.05). MRD≥10-4 at the end of induction therapy is an independent risk factor for poor prognosis (hazard ratio of event-free, 9.101 [95% CI 2.03-40.86], P =0·004, compared to those with a MRD of <10-4). Conclusion: Pediatric ALL patients with PDGFRB fusions have a poor outcome. This subtype of ALL is associated with high-risk clinical features such as older age, high WBC count at diagnosis, high MRD after induction therapy, and increased risk of leukemia relapse. The addition of TKI to chemotherapy can improve the early remission rate of treatment, but does not improve the long-term survival. Profitably, the TKI treatment can provide an opportunity for subsequent treatment, including HSCT. Additionally, HSCT has indeed potential to reduce the relapse rate for this subtype patients. HSCT is recommended for this subtype patients who achieve CR1. The MRD≥10-4 or more at the end of induction therapy was independently predictive factor for unfavorable outcome and stratified treatment based on MRD may improve survival of these patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

ALL-257 Detection of Deletion in the IKZF1 Gene and the NOTCH1 Signaling Pathway in Patients With T-Cell Acute Lymphoblastic Leukemia: Data of the RALL Study Group

Mutation in the NOTCH1 is the most frequent event in patients with T-acute lymphoblastic leukemia (ALL). This mutation is associated with a favorable prognosis. Deletion of the IKZF1 gene, occurring in patients with T-ALL in about 10% of cases, is a prognostically unfavorable anomaly. To evaluate the survival of patients with T-ALL depending on the immunophenotype, as well as to analyze the frequency of occurrence of anomalies in the NOTCH1 and IKZF1 genes. The analysis included 91 patients with T-ALL/LBL receiving therapy according to the ALL-2016 protocol. The immunophenotype was determined by flow cytofluorimetry. Anomalies in the IKZF1 and NOTCH1 were investigated by fragment analysis according to Caye A, et al and Campregher PV, et al in 25 and 29 patients, respectively. From December 2016 to June 2021, 91 patients with T-ALL were included in the ALL-2016 protocol (64 men and 27 women, median 32 years: 6-TIV, 11-ETP-ALL, 32-TI/II, 42-TIII). 3-year overall survival in the ETP group was 28%, TI/II-50%, TIII-73% and TIV-83%. Molecular anomalies in NOTCH1 were analyzed by fragment analysis in 29 patients (1-undifferentiated variant, 8-ETP-ALL, 9-TIII, 10-TIII, 1-TIV), in IKZF1 genes in 25 patients (1-undifferentiated variant, 5-ETP-ALL, 8-TIII, 10-TIII, 1-TIV). None of the patients had deletions in the IKZF1. Anomalies in the NOTCH1 were detected in 19 (65.5%) patients (4-ETP-ALL, 7-TII, 7-TIII, 1-TIV). At the same time, 15 of them (51%) had NOTCH1 deletions, 4 (14%) had insertions. 5 patients without abnormalities in the NOTCH1 (50%) died as a result of a resistant or relapse, 5 patients are alive in remission. In the group of patients with an anomaly in the NOTCH1, 3 patients (16%) died, the remaining 16 patients (84%) are alive in remission. Overall survival was 84% in the group with anomalies versus 50% in the group without anomalies (p=0.05). Anomalies in the NOTCH1 are the most common in patients with T-ALL. This mutation is associated with a more favorable prognosis.

Myeloid checkpoint blockade improves killing of T-acute lymphoblastic leukemia cells by an IgA2 variant of daratumumab.

Antibody-based immunotherapy is increasingly employed to treat acute lymphoblastic leukemia (ALL) patients. Many T-ALL cells express CD38 on their surface, which can be targeted by the CD38 antibody daratumumab (DARA), approved for the treatment of multiple myeloma. Tumor cell killing by myeloid cells is relevant for the efficacy of many therapeutic antibodies and can be more efficacious with human IgA than with IgG antibodies. This is demonstrated here by investigating antibody-dependent cellular phagocytosis (ADCP) by macrophages and antibody-dependent cell-mediated cytotoxicity (ADCC) by polymorphonuclear (PMN) cells using DARA (human IgG1) and an IgA2 isotype switch variant (DARA-IgA2) against T-ALL cell lines and primary patient-derived tumor cells. ADCP and ADCC are negatively regulated by interactions between CD47 on tumor cells and signal regulatory protein alpha (SIRPα) on effector cells. In order to investigate the impact of this myeloid checkpoint on T-ALL cell killing, CD47 and glutaminyl-peptide cyclotransferase like (QPCTL) knock-out T-ALL cells were employed. QPTCL is an enzymatic posttranslational modifier of CD47 activity, which can be targeted by small molecule inhibitors. Additionally, we used an IgG2σ variant of the CD47 blocking antibody magrolimab, which is in advanced clinical development. Moreover, treatment of T-ALL cells with all-trans retinoic acid (ATRA) increased CD38 expression leading to further enhanced ADCP and ADCC, particularly when DARA-IgA2 was applied. These studies demonstrate that myeloid checkpoint blockade in combination with IgA2 variants of CD38 antibodies deserves further evaluation for T-ALL immunotherapy.