Positive Leukemia Research Articles

Frontline combination of ponatinib and hyper-CVAD in Philadelphia chromosome-positive acute lymphoblastic leukemia: 80-months follow-up results.

The combination of ponatinib, a third-generation BCR::ABL1 tyrosine kinase inhibitor, with hyper-CVAD chemotherapy resulted in high rates of complete molecular remissions and survival, without the need for stem cell transplantation (SCT) in most patients with Philadelphia chromosome(Ph)-positive acute lymphocytic leukemia (ALL). Confirming these results in a large cohort of patients with longer follow-up would establish this regimen as a new standard of care. Adults with newly diagnosed Ph-positive ALL were treated with the hyper-CVAD regimen. Ponatinib was added as 45 mg daily × 14 during induction, then 45 mg daily continuously (first 37 patients) or 30 mg daily continuously, with dose reduction to 15 mg daily upon achievement of a complete molecular response (CMR; absence of a detectable BCR::ABL1 transcript by quantitative reverse transcription polymerase-chain reaction at a sensitivity of 0.01%). Maintenance therapy consisted of daily ponatinib and vincristine-prednisone monthly for 2 years, followed by daily ponatinib indefinitely. Twelve intrathecal injections of cytarabine alternating with methotrexate were given as central nervous system prophylaxis. The trial is registered on clinicaltrials.gov with the identifier NCT01424982. Eighty-six patients were treated. Their median age was 46 years (range, 21-80). All 68 patients with active disease at the initiation of therapy achieved complete response (CR) The cumulative CMR rate was 86%. Twenty- patients (23%) underwent allogeneic SCT. With a median follow-up of 80 months (range, 16-129 months), the estimated 6-year event-free survival rate was 65% and the overall survival rate was 75%. There was no difference in outcome by performance of allogeneic SCT in first CR. Common grade 3-5 adverse events included infection (n= 80, 93%), increased liver transaminases (n= 26, 31%) and total bilirubin (n= 13, 15%), hypertension (n= 15, 17%), pancreatitis (n= 13, 15%), hemorrhage (n= 12, 13%), and skin rash (n= 9, 10%). Two ponatinib-related deaths from myocardial infarction (3%; at months 2.6 and 4.3, respectively; both in CR) in the first 37 patients treated led to the ponatinib dose-modifications mentioned earlier, with no further ponatinib-related deaths observed. The long-term results of ponatinib and hyper-CVAD continue to demonstrate excellent outcome results and acceptable safety data, indicating that this strategy is another standard of care approach in frontline Ph-positive ALL.

Positive Myeloproliferactive Dissorder and Chronic

Myeloproliferative neoplasms (MPN) arise from hematopoietic stem cells with somatically altered tyrosine kinase signaling. The World Health Organization (WHO) classifies them according to the presence of the BCR/ABL translocation in BCR/ABL positive chronic myelogenous leukemia (CML) and those which do not express BCR/ABL including; polycythemia vera, essential thrombocytosis and primary myelofibrosis. This last group is characterized by the expression of other mutations such as JAK2, MPL and CALR. JAK2, the most common, occurs in approximately 50% of cases (1, 2). According to literature, BCR/ABL and JAK2 are mutually exclusive (1, 2), however in some cases they can coexist. Two hypotheses have been postulated. The first one proposes that exists two cell clones, exhibiting each one, one mutation. Under treatment pressure of the dominant clone, the other one becomes over expressed. The second hypothesis states that exists one cell clone that simultaneously expresses both mutations (3, 4).

PROFILE OF CD150 EXPRESSION IN BONE MARROW CELLS OF PATIENTS WITH ACUTE MYELOID LEUKEMIA.

Acute myeloid leukemia (AML) is a highly heterogeneous disease accompanied by thearrest of myeloid cell lineage differenti-ation due to accumulation of genetic abnormalities and clonal proliferation of myeloid blasts. Finding thedifferentially expressed molecules and studying their function within AML subgroups may help to improve diagnosis and prognosis with theaim of developing selected therapies for AML subsets. Theaim of this study was to reveal theprofile of CD150cell surface expression on bone marrow (BM) cells of AML patients. Thestudy was performed on samples of BM aspirates from 55patients with primarily diagnosed AML. Flow cytometry analysis was applied for theevaluation of immunophenotype profile and CD150cell surface expression on BM cells from AML patients. Four AML subtypes (M1, M2, M3and M5) were identified. TheCD150expression was found in 14 (25.5%) cases predominantly of AML M3subtype. CD150expression was detected on 43.2-83.8% of leukemia cells in AML M3. Thefrequency of CD150positive cases of non-M3AML subtypes was low: all AML M1cases were CD150-negative, while only 1 (10.0%) of 10patients with AML M2and 6 (19.4%) of 31patients with AML M5were CD150positive. Themedian percentage of CD150positive leukemia cells and theindex of mean fluorescence intensity in AML M3cases were significantly higher than in non-M3AML cases (p < 0.05). TheCD150expression was significantly associated with CD11c, CD11b, CD14, CD34, CD36, CD56and HLA-DR negative expression and CD33, CD38, CD117positive expression among theexamined cohort of patients with AML M3. High level of CD150expression is a unique feature of AML M3subtype and may serve as additional phenotype marker for theidentification of blast cells with impaired maturation at thepromyelocyte stage and thedevelopment of AML M3. At thesame time, therevealed negative association of CD150expression with poor prognostic factor CD56in AML M3subtype also allows us to suggest potential prognostic value of CD150examination in AML patients.

Novel modular chimeric antigen receptor spacer for T cells derived from signal regulatory protein alpha Ig-like domains.

Background: T cells equipped with chimeric antigen receptors (CAR) have shown remarkable efficacy in targeting B lineage malignancies. Improvement of the CAR structure is needed, however, with a view to developing flexibly modifiable spacers that are inert in interactions with unwanted cells. Specifically, binding to cells carrying receptors for IgG's crystallizable fragment (FcR), that recognize IgG-derived domains in CARs is to be avoided. Methods: Two novel CARs targeting the CD19 antigen where the IgG1-CH2 and -CH3 domains were replaced with Ig-like domains from signal-regulatory protein α (SIRPα) were designed in silico. An IgG1-based CAR and a CAR lacking both SIRPα and IgG1 domains were used as comparators. The phenotype and memory phenotype of the expanded cells were analyzed by flow cytometry, and CAR T cell activation and cytotoxic efficacy were assessed in co-culture experiments in response to CD19+ target cells. Unwanted interactions with FcR-expressing myeloid cells were interrogated in co-culture assays with THP-1 monocytic cells. Results: T cells carrying the novel SIRPα-based CARs enacted potent in vitro cytotoxicity against CD19 positive B-lineage leukemia cells, comparable to traditional IgG1-based CAR T cells. Co-culture of IgG1-based CAR T cells with FcR-expressing THP-1 monocytic cells led to prominent cell surface expression of CD69 on T cells together with production of Interleukin (IL)-2 and Interferon-γ, and production of IL-1β, indicating activation of the T cells and monocytes, respectively. Longer co-culture led to killing of the monocytes. No signs of T cell nor monocyte activation were detected in co-cultures of SIRPα-based CAR T cells with THP-1 cells. Arming T cells with the SIRPα-based CARs favored differentiation towards CD4+ phenotype during expansion, while the effects on memory phenotype of the T cells were equivalent between the SIRPα- and IgG1-based CARs. In a pilot experiment, T cells modified with one of the SIRPα-based CARs showed dose dependent leukemia cell control. Conclusion: The novel SIRPα based spacers offer a suitable backbone for developing chimeric antigen receptors that evade the off-target binding to FcR while the cells retain a favorable memory phenotype and efficient cytotoxicity, establishing a promising candidate for future in vivo and clinical testing.

A key role of the WEE1-CDK1 axis in mediating TKI-therapy resistance in FLT3-ITD positive acute myeloid leukemia patients

The insertion site of the internal tandem duplications (ITDs) in the FLT3 gene affects the sensitivity to tyrosine kinase inhibitors (TKIs) therapy in acute myeloid leukemia (AML). Patients with the ITD in the tyrosine kinase domain lack effective therapeutic options. Here, to identify genotype-driven strategies increasing the TKI therapy efficacy, we developed SignalingProfiler, a strategy supporting the integration of high-sensitive mass spectrometry-based (phospho)proteomics, RNA sequencing datasets with literature-derived signaling networks. The approach generated FLT3-ITD genotype-specific predictive models and revealed a conserved role of the WEE1-CDK1 axis in TKIs resistance. Remarkably, pharmacological inhibition of the WEE1 kinase synergizes and strengthens the pro-apoptotic effect of TKIs therapy in cell lines and patient-derived primary blasts. Finally, we propose a new molecular mechanism of TKIs resistance in AML and suggest the combination of WEE1 inhibitor and TKI as a therapeutic option to improve patients clinical outcome.

The effect of milk from cows infected with bovine leukemia virus on tumor development in Balb/c mice

The article describes studies on the effect of milk from immunodiffusion test - positive leukemia cows on the dynamics of tumor development in BALB/c mice caused by syngeneic plasmacytoma (myeloma) Sp.2/0 - Ag.8 cell line. The research results have shown that raw milk from cows infected with bovine leukemia virus significantly increases the rate of tumor development in both male and female mice. Boiling of that milk does not reduce its negative effect on the organism of animals. Female mice are more susceptible to milk from cows infected with bovine leukemia virus. After including milk from cows infected with the bovine leukemia virus into the diet of animals, a significant difference in tumor size from the control group of mice was manifested in both males and females, but the volumes of tumors in females were an order of magnitude larger.

Case report: A rare case of TBL1XR1-RARB positive acute promyelocytic leukemia in child and review of the literature.

Some forms of acute myelogenous leukemia (AML) share typical morphological and immunophenotypic features of acute promyelocytic leukemia (APL) but are negative for promyelocytic leukemia-retinoic acid receptor alpha (PML-RARA) fusion. These forms of AML are known as variant APL. Some variants of APL present with retinoic acid receptor beta (RARB) fused or rearranged with partner genes. RARB-positive APL is very rare, resistant to all-trans retinoic acid (ATRA), and associated with poor prognosis. Here, we reported one case with TBL1XR1-RARB positive APL, featured by early onset and no apparent bleeding tendency or coagulation dysfunction. This patient was resistant to ATRA and arsenic trioxide (ATO), but was good responsive to conventional chemotherapy for AML. The case report was followed by a literature review.

A Phase II Study of the Combination of Decitabine, Venetoclax and Ponatinib in Patients with Chronic Myeloid Leukemia (CML) in Myeloid Blast Phase (MBP) or Philadelphia-Chromosome Positive (Ph+) Acute Myeloid Leukemia (AML)

Background: Advanced phases of CML and Ph+ AML are therapeutic challenges. Preclinical and early clinical data have shown that combining venetoclax (VEN) with a tyrosine kinase inhibitor (TKI) works synergistically in Ph+ myeloid leukemias (Leonard et. al., Sci Transl Med 2016 and Maiti et. al., Acta Haematologica 2020). We hypothesized that combining a potent TKI (ponatinib) with a hypomethylating agent plus VEN backbone might lead to high rates of response that could facilitate potentially curative allogeneic stem cell transplantation (SCT). Methods: This is a single-arm phase II study (NCT04188405) that enrolled patients (pts) ≥ 18 years (yrs) of age with newly diagnosed or relapsed/refractory advanced phase CML [CML in accelerated phase (CML-AP) or CML-MBP] or Ph+ AML. Pts were required to have an ECOG PS ≤ 3 and adequate liver and renal function. Pts with clinically significant cardiovascular comorbidities, including recent or active myocardial infarction, heart failure, clinically significant arrhythmias, cerebrovascular accident, venous thromboembolism or uncontrolled hypertension were excluded. Pts were treated with a combination of decitabine (DAC) (20 mg/m2/day) x 5 days, VEN (400 mg equivalent dose/day) x 21 days, and ponatinib (45mg/day) x 21 days in cycle 1 and then continuously in cycle 2+ (Fig. 1A). Ponatinib was reduced to 30mg/day for pts in CR/CRi and to 15 mg/day for pts in CMR. For C1 only, pts who had not received ponatinib prior to trial initiation had a lead-in phase of ponatinib monotherapy for 7 days before the initiation of DAC and VEN. Cycles were repeated every 28 days, as permitted by peripheral blood count recovery, and pts could receive a maximum of 24 cycles. Intrathecal prophylaxis with 4 doses of cytarabine was recommended. The primary objective was to determine the overall response rate of the regimen. Results: From July 2020 to May 2022, 12 pts were treated, 1 pt with de novo Ph+ AML and 11 pts with advanced phase CML (2 pts with CML-AP and 9 pts with CML-MBP). The median age of the pts was 45 yrs (range, 28-71 yrs). Three pts (25%) had received no prior therapy (1 each with Ph+ AML, CML-AP and CML-MBP). Nine pts (75%) had prior TKI exposure (including prior ponatinib exposure in 4 pts) and 3 pts (25%) had received prior chemotherapy. Eleven pts (92%) had the p210 transcript of BCR::ABL1. Eight pts (67%) had additional cytogenetic abnormalities, including 5 pts with complex karyotype and 2 pts with a MECOM rearrangement. 4 of 11 tested pts (36%) had an ABL1 kinase domain mutation detected at enrollment (T315I, Q252H, L384M and E255K in 1 pt each). The median number of cycles received was 2 (range, 1-7). Nine pts (75%) responded, including 1 with CR, 3 with CRi and 5 with MLFS as best response. The CR/CRi rate was 33%. Best response was achieved after cycle 1 in 7 of 9 responders. Both pts with CML-AP responded, and 2 of 3 pts without prior TKI exposure responded. One of the responders achieved an MMR. After a median follow-up of 9.1 months, the median overall survival (OS) is 9.5 months, and the estimated 9-month OS is 64%. The median relapse-free survival 5.1 months. Among the 9 responders, 3 pts are in continuous response without SCT, 1 pt proceeded to SCT and is still alive, 3 pts did not undergo SCT and relapsed (2 of whom had a complex karyotype), 1 pt died in MLFS, and 1 pt was lost to follow-up (Fig. 1B). The one pt who proceeded to SCT achieved MLFS as best response pre-SCT and remains in MRD-negative CR 22 months post-SCT. The toxicity profile of the regimen was similar to historical expectations with DAC plus VEN. One pt developed grade 3 mucositis, which was considered related to study therapy. No grade 4-5 related non-hematological adverse events were observed. Related grade 1-2 non-hematological adverse events included rash in 2 pts, and transaminitis, creatinine elevation and mucositis in 1 pt each. Of the 8 pts who received > 1 cycle of study therapy, 4 pts had delay (>1 week) in initiation of a subsequent treatment cycle due to cytopenias; for 2 of these pts the duration of VEN was reduced. The 60-day mortality rate was 0%. Two pts died on the study, one from leukemia and one from infection while in MLFS. Conclusion: The triplet combination of DAC, VEN and ponatinib was well-tolerated in pts with advanced Ph+ myeloid leukemias. In this poor-risk population, a marrow remission rate of 75% was achieved, although durations of remission were modest in the absence of consolidative SCT. This study continues to accrue pts. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Outcomes of Olverembatinib Based Regimen in Patients with Philadelphia Chromosome-Positive Advanced Leukemias: A Real-World, Single-Center Experience

Background Philadelphia chromosome-positive (Ph+) advanced leukemias including chronic myeloid leukemia blast phase and acute lymphoblastic leukemia have poor outcomes. Olverembatinib, a novel BCR-ABL tyrosine kinase inhibitor, has anti-tumor activity in Philadelphia chromosome-positive (Ph+) chronic myeloid leukemia cells(CML) and tumor models, is currently approved for treating resistant CML patients with T315I mutation by the National Medical Products Administration in China, and in phase Ib clinical trials in the US for ponatinib resistant and/or intolerant CML patients. Olverembatinib also inhibited p-KIT, p-AKT, p-ERK1/2, and p-STAT3 to a higher extent than ponatinib, and suppresses pre-B ALL cells through inhibition of the SRC kinase and PI3K/AKT pathways and may be a potential therapeutic agent for the management of pre-B ALL. However, the efficacy of Olverembatinib based regimen in Philadelphia chromosome- positive advanced leukemias is still unknown. Methods We conducted a retrospective study on patients with Philadelphia Chromosome- Positive Advanced Leukemias including Ph+ CML (n=11) and Ph+ ALL (n=5) who received a Olverembatinib-based regimens in our center. Results The trial included 11 patients with Ph+ CML and 5 patients with Ph+ ALL who had relapsed after transplantation. Among the 11 CML patients, there were 2 CML-AP, 6 chronic myeloid leukemia-myeloid blast phase (MBP), 2 chronic myeloid leukemia-lymphoblastic bast phase(LBP), 1 chronic myeloid leukemia- myeloid and lymphoblastic mixed blast phase. Median age was 34 years and the median follow-up time was 4 months. Also, two (18.2%) CML-AP patients were treated witholverembatinib alone. Six (54.5%) CML-BP patients were treated with olverembatinib in combination with demethylating agents. And two patients with LBP received VDP and stem cell transplantation (SCT)-based regimens combined with olverembatinib. Treatment for patients with chronic myeloid leukemia- myeloid and lymphoblastic mixed blast phase is consistent with patients with MBP. The overall response rate of 8 evaluable patients was 100%. CR/CRi rate was 100%, and CCyR/PCyR rates 62.5% vs 37.5%, respectively. In addition, 6 patients achieved major molecular responses (MMR). the remaining three patients were not yet due for evaluation. Of 5 patients with ph+ ALL that relapsed after transplantation, 1patient was treated with blinatumomab and olverembatinib, 2 patients were treated with intensive chemotherapy and olverembatinib followed by donor lymphocyte infusion(DLI), The treatment for the remaining 2 patients was Anti-CD19 chimeric antigen receptor T cell (CAR-T) technology followed by olverembatinib, and this 5 Ph+ ALL patients achieved complete molecular remission(CMR). In addition, treatment-related adverse reactions included cytopenias, skin pigmentation, proteinuria and high triglycerides, all grades less than 2. No deaths have occurred. Conclusion Olverembatinib-based regimens show encouraging activity and safety in very heavily pre-treated, advanced Ph+ leukemias.

Efficacy and Safety of Vodobatinib in Patients (pts) with Chronic Phase Philadelphia Positive Chronic Myeloid Leukemia (Ph+ CML): A Sub Group Analysis By Lines of Tyrosine Kinase Inhibitor (TKI) Therapy

Efficacy and Safety of Vodobatinib in Patients (pts) with Chronic Phase Philadelphia Positive Chronic Myeloid Leukemia (Ph+ CML): A Sub Group Analysis By Lines of Tyrosine Kinase Inhibitor (TKI) Therapy

Utilizing CLL-1 Positive Leukemia Stem Cells As a Marker of Early Treatment Response in Acute Myeloid Leukemia

Background: Current methods to evaluate therapeutic response in Acute Myeloid leukemia (AML) rely primarily on morphological evaluation of leukemia blasts in bone marrow at end of treatment cycles, with complete remission (CR) defined as less than 5% bone marrow blast counts without leukemic phenotype. Measurable residual disease (MRD) has been shown highly prognostic for outcome. MRD by molecular methods is limited to subgroup of patients with defined genetic mutations. MRD detection by Multicolor Flow Cytometry (MFC) is still being standardized. Leukemia stem cells (LSC) has been established as source of treatment resistance and disease recurrence in AML. In this study, we aim to analyze change in LSC during treatment using immunophenotypically defined markers and examine whether information gained from LSC analysis could provide additional insights into disease course and assist management of AML under routine clinical practice setting. Methods: We analyzed flow cytometry data from bone marrow aspirate (BMA) and/or peripheral blood (PB) in patients with AML (excluding acute promyelocytic leukemia) before and after treatment. "Leukemic blasts” were identified by morphology and Leukemia-associated Immunophenotype by MFC. Hematopoietic stem cells (HSCs) gating was based on CD45dim/SSC and CD34+CD38low/- expression. Leukemic stem cells (LSC) fractional change was analyzed using CD371 (CLL-1), CD366 (TIM3), and CD45RA percent of positive over HSCs. A chart review was conducted to determine correlation between LSC fractional change and clinical outcome. Statistical analysis was performed using GraphPad Prism version 9. Results: Fifty AML cases with both pre- and post-treatment data from a single institution in the past 3 years were included in this study. Consistent with published literature, patients with significant reduction in LSC subsets was associated with longer remission, while patients with no significant decrease in the LSC subsets demonstrated no or only a temporary remission. Intriguingly, several patients with drastic reduction in LSC after induction chemotherapy achieved complete remission, despite initial post-treatment bone marrow showing persistence of ">5% leukemia blasts” indicating possibly failed induction therapy. In contrast, in patients with persistently high or increased LSC fraction post-treatment, overt clinical relapse occurred quickly despite "absence” or <1% of leukemia blast differential counts in initial bone marrow. In a series of patients who received allogeneic stem cell transplant (Allo-SCT), rise of LSC fraction post Allo-SCT was associated with early AML relapse (within 6 months), while the decrease of LSC fraction was associated with prolonged remission (Representative patient cases shown in Figure 1). Lastly, among the three LSC-specific markers analyzed, the changes in LSC characterized by CLL-1 expression showed the best correlation with clinical course. In addition, the LSC fraction measured by CLL-1 in the peripheral blood is highly correlated with that of the bone marrow (R2=0.93, P<0.0001). Conclusion: This retrospective study explored the value and applicability of LSC detection in routine clinical practice. Importantly, conventional methods relying on evaluating leukemic blast counts in post-treatment bone marrow can sometimes over- or under- estimate the benefit of certain treatment regimen, leading to inappropriate continuation of the same treatment or switch of a potentially effective treatment regimen. In those groups of patients, the change in LSC fraction provides additional information for early treatment response evaluation, refines outcome prediction, and helps with treatment regimen selection. Trending LSC fractional change identifies impending relapse in post Allo-SCT. LSC fractional change determined by CLL-1 showed the best clinical correlation and should be incorporated into routine evaluation for AML patient in clinical practice, and possibly for those participating in investigational trials. Future studies will focus on collecting more data to validate above findings, determining the thresholds of LSC fractional change which constitute potential treatment response vs failure and testing the feasibility of tracking LSC subsets non-invasively in peripheral blood during treatment to predict therapeutic outcome and potentially guide treatment decisions. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Whole Genome Sequencing for Detection of Prognostically Relevant Genetic Abnormalities in Paediatric Acute Myeloid Leukaemia

Background Genetics is important in the diagnosis of paediatric acute myeloid leukaemia (AML). A number of chromosomal abnormalities and mutations have prognostic significance and timely genetic analysis is essential for risk stratification. The recent Myechild 01 clinical trial for children with AML required testing for >20 different genetic abnormalities to assign newly diagnosed patients into three prognostic genetic risk groups: good, intermediate and poor (Figure A). Throughout the trial a combination of techniques was employed to detect these abnormalities; including chromosomal analysis, FISH for cytogenetically cryptic abnormalities, notably the poor risk fusions: t(5;11)(q35;p15)/NUP98::NSD1, inv(16)(p13q24)/CBFA2T3::GLIS2, t(7;12)(q36;p13)/ETV6::MNX1, as well as molecular testing for NPM1, FLT3-ITD and CEBPA mutations. Here, we present a small retrospective study demonstrating that whole genome sequencing (WGS) has the potential to detect all prognostically significant abnormalities in childhood AML within a single test. Methods WGS was performed on matched diagnostic and remission samples from 25 paediatric AML patients entered to the Myechild 01 treatment trial. Remission samples, minimal residual disease negative by flow cytometry and/or molecular testing, were used as germline controls. For validation, whole transcriptome sequencing (WTS) was performed on diagnostic RNA from each patient in the study. Results Leukaemia associated genetic abnormalities were detected in all 25 patients, including structural variants associated with gene fusions (n=14), single nucleotide variants (SNV) (n=20) and/or copy number abnormalities/aneuploidies (n=17). All gene fusions and mutations were confirmed by WTS. WGS reliably detected all risk stratifying abnormalities identified by standard techniques (Figure B), including 11 patients with gene fusions: RUNX1::RUNX1T1 (n=6), CBFB::MYH11 (n=3), KMT2A::MLLT3 (n=1) and KMT2A::MLLT10 (n=1) and 3 patients with copy number abnormalities: del(5q) (n=2) and del(12p) (n=1) and one patient with an NPM1 mutation. Among 10 patients with no risk stratifying abnormalities detected by standard techniques, WGS identified risk defining genetic features in 3 of them. Two patients could be reclassified as good risk due to the discovery of biallelic CEBPA mutations. Interestingly, one of the CEBPA positive patients presented with a homozygous mutation (VAF=0.97), which WGS revealed to be the consequence of copy number neutral loss of heterozygosity for the long arm of chromosome 19. A single patient could be reclassified as poor risk due to the finding of a rearrangement of MECOM with CDK6. Cytogenetics showed a deletion of 7q as the sole karyotypic abnormality, which was revealed to be an unbalanced der(7)t(3;7)(q21.2;q26.2) translocation by WGS. Over-expression of MECOM was confirmed by WTS. This patient also had a GATA2 mutation. A risk defining abnormality was not detected by either standard testing or WGS in 7 patients, however 5 of them presented with chromosomal abnormalities which were confirmed and further characterised by WGS, including aneuploidies (n=3) and/or structural rearrangements (n=4). Across the cohort, WGS detected a wide range of abnormalities not currently used in the risk stratification algorithm, including the fusions genes: NPM1::MLF1 (n=1) and PICALM::MLLT10 (n=2), which, although not stratifying in Myechild 01, have been reported to be associated with a good and a poor prognosis, respectively. SNV in genes commonly mutated in AML were observed in 19 patients (76%) including NRAS (n=6), KIT (n=3) EZH2 (n=3) and GATA2 (n=2). It has been suggested that the co-occurrence of mutations may be used to refine risk groups in childhood AML, notably the presence of KIT and NRAS mutations in RUNX1::RUNXT1 and CBFB::MYH11 positive leukaemia, respectively. Conclusion This small pilot study has demonstrated the ability of WGS to identify a wide range of genetic abnormalities that are currently used for risk stratification in childhood AML. The full spectrum of genomic events observed here highlights the potential of WGS to refine risk stratification and identify novel genetic changes as therapeutic targets in future trials. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Abstract 14022: Involvement of Rho-Associated Coiled-Coil Containing Kinase (ROCK) in BCR-ABL1 Tyrosine Kinase Inhibitor Cardiovascular Toxicity

Introduction: Second and third generation BCR-ABL1 tyrosine kinase inhibitors (TKIs) are associated with cardiovascular adverse events (CVAE) in patients with Philadelphia-chromosome positive (Ph+) leukemia. Hypothesis: We hypothesized that second and third generation BCR-ABL1 TKIs may cause CVAE through activation of Rho-associated protein kinase (ROCK). Methods: Peripheral blood mononuclear cells from 53 Ph+ patients on TKIs and 15 control patients without Ph+ leukemia were assessed for ROCK activity through capillary electrophoresis (median follow-up = 26 months [Q1-Q3, 5-37]). We also investigated the effects of TKIs and ROCK on endothelial dysfunction in vitro , which could contribute to CVAE. Results: Patients receiving second and third generation TKIs had 1.6-fold greater ROCK activity compared to both patients receiving imatinib and control patients. Elevated ROCK activity was associated with increased incidence of CVAE in Ph+ leukemia patients (Figure). In endothelial cells in vitro , we found that dasatinib and ponatinib treatment lead to changes in actin intensity and endothelial permeability, which can be reversed by pharmacologic inhibition of ROCK. Ponatinib leads to decreased cell proliferation, but this was not accompanied by senescence. Dasatinib and ponatinib treatment lead to phospho-inhibition of endothelial nitric oxide synthase (eNOS) and decreased nitric oxide production. ROCK inhibition reversed endothelial permeability and eNOS-related endothelial dysfunction. Imatinib and nilotinib induce phosphorylation of p190RhoGAP. Conclusions: Our findings suggest ROCK activity may be a prognostic indicator of CVAE in patients receiving BCR-ABL1 TKIs. ROCK inhibition may be a promising approach in reducing the incidence of CVAE associated with second and third generation BCR-ABL1 TKIs.

Abstract 14372: Case Series: Non-Lipid Plaque Formation as the Mechanism for Ponatinib-Induced Vascular Toxicity Using Operator-Assisted Virtual Histology Optical Coherence Tomography

Introduction: The third-generation tyrosine kinase inhibitor ponatinib approved for treating chronic myeloid leukemia (CML) rendered drug resistant with a threonine-315-isoleucine (T315I) mutation is associated with increased incidence of vascular adverse events (VAEs). This case series sought to elucidate the mechanism of these VAEs. Methods: Two patients with CML and two patients with Philadelphia chromosome positive acute lymphocytic leukemia (Ph+ ALL) on at least one year of ponatinib therapy were imaged with optical coherence tomography (OCT) after presenting with angina. An additional four patients from an institutional OCT database were matched across clinical parameters (age, sex, diabetes, prior coronary artery disease (CAD) history, prior congestive heart failure (CHF) history, hypertension, and family history of CAD) to serve as controls. Plaque composition was assessed using operator-assisted virtual histology OCT (vOCT). Results: Clinical presentations included 2 cases of NSTEMI, 1 case of unstable angina, and 1 case of stable angina pectoris among the 4 ponatinib patients. Atherosclerosis was composed primarily of calcium and fibrous tissue with minimal lipid presence (0 ± 0°) compared to the matched controls. The matched controls had a mean non-zero lipid content of 48.43 ± 18.36°, ( P &lt; 0.05) (Table 1). Three of the four patients had cardiovascular risk factors. Conclusions: Ponatinib causes VAEs through non-lipid plaque formation driving a symptomatology consistent with an oxygen supply-demand mismatch. This mechanism is distinct from traditional pathways of excess lipid and plaque instability causing coronary artery disease and merits further investigation.

Understanding a high-risk acute myeloid leukemia by analyzing the interactome of its major driver mutation.

The WHO classifies t(6;9)-positive acute myeloid leukemia (AML) as a subgroup of high-risk AML because of its clinical and biological peculiarities, such as young age and therapy resistance. t(6;9) encodes the DEK/NUP214 fusion oncoprotein that targets only a small subpopulation of bone marrow progenitors for leukemic transformation. This distinguishes DEK/NUP214 from other fusion oncoproteins, such as PML/RARα, RUNX1/ETO, or MLL/AF9, which have a broad target population they block differentiation and increase stem cell capacity. A common theme among most leukemogenic fusion proteins is their aberrant localization compared to their wild-type counterparts. Although the actual consequences are widely unknown, it seems to contribute to leukemogenesis most likely by a sequester of interaction partners. Thus, we applied a global approach to studying the consequences of the aberrant localization of t(6;9)-DEK/NUP214 for its interactome. This study aimed to disclose the role of localization of DEK/NUP214 and the related sequester of proteins interacting with DEK/NUP214 for the determination of the biology of t(6;9)-AML. Here we show the complexity of the biological consequences of the expression of DEK/NUP214 by an in-depth bioinformatic analysis of the interactome of DEK/NUP214 and its biologically dead mutants. DEK/NUP214's interactome points to an essential role for aberrant RNA-regulation and aberrant regulation of apoptosis and leukocyte activation as a significant determinant of the phenotype of t(6;9)-AML. Taken together, we provide evidence that the interactome contributes to the aberrant biology of an oncoprotein, providing opportunities for developing novel targeted therapy approaches.

Blinatumomab plus hyper-CVAD: the prelude to a new era in acute lymphocytic leukaemia

Blinatumomab, a CD19/CD3 bispecific T-cell engager active as monotherapy in relapsed or refractory B-cell acute lymphocytic leukaemia, was approved for adults with positive minimal residual disease (MRD) in Europe and the USA in 2018. Approval was granted on the basis of a single-arm, phase 2 study, in which 78% of included patients had an MRD-negative complete response and promising outcomes and acceptable toxicity were seen.1,2 Early experiences have also been encouraging with blinatumomab as first-line treatment of Philadelphia chromosome (Ph)-positive and Ph-negative acute lymphocytic leukaemia in older adults (aged >65 years)—a population that has previously had poor outcomes owing to adverse biology and poor tolerance to chemotherapy.

A Dasatinib-Induced Chylothorax Persisting After the Discontinuation of Dasatinib

Dasatinib is a rare cause of chylothorax.Chylothorax should be included in the differential diagnosis in patients developing pleural effusions when treated with dasatinib.The cornerstone of the treatment is tapering the dose or discontinuing the dasatinib therapy and changing it to another tyrosine kinase inhibitor.

Asciminib (Scemblix): A third-line treatment option for chronic myeloid leukemia in chronic phase with or without T315I mutation.

To review the pharmacology, efficacy, safety, dosing and administration, and place in therapy of asciminib, an oral tyrosine kinase inhibitor (TKI) used as a third-line treatment option for Philadelphia chromosome-positive chronic myeloid leukemia (CML) in chronic phase. CML is a rare cancer caused by a chromosomal translocation that forms a fusion of the BCR and ABL1 genes on chromosomes 22 and 9. Until recently, patients for whom first-line treatment options failed were treated with TKIs that bind to the adenosine triphosphate-binding site on BCR-ABL1. However, because of similar mechanisms of action, there continues to be an unmet need in patients for whom at least 2 TKIs have failed or those with a T315I mutation unable to tolerate ponatinib. In October 2021, the Food and Drug Administration approved asciminib (Scemblix), the first TKI specifically targeting the ABL1 myristoyl pocket (STAMP) via allosteric binding, as a third-line option for patients with chronic-phase (CP)-CML. Asciminib received accelerated approval due to meeting its primary endpoint at week 24, demonstrating a major molecular response rate of 25.5% for patients on asciminib compared to 13.2% for those receiving bosutinib. In addition, patients on asciminib achieved a higher rate of complete cytogenetic response at 40.8% compared to a rate of 24.2% for bosutinib. Clinicians prescribing asciminib should monitor for increased levels of pancreatic enzymes, hypertension, cardiovascular toxicity including ischemic and thromboembolic conditions, and decreased numbers of neutrophils and platelets, as these may require treatment interruption, dose reduction, or treatment discontinuation. Asciminib is a unique targeted TKI that provides clinicians with an additional third-line and beyond treatment option for adults with CP-CML regardless of mutation status as well as a second TKI treatment option for patients harboring a T315I mutation.

AML-029 Quizartinib Prolonged Overall Survival (OS) vs Placebo Plus Intensive Induction and Consolidation Therapy Followed by Single-Agent Continuation in Patients Aged 18-75 Years With Newly Diagnosed FLT3–Internal Tandem Duplication Positive (FLT3-ITD+) Acute Myeloid Leukemia (AML)

Quizartinib is an oral, potent, selective, type II FLT3 inhibitor with prolonged OS as a single-agent in relapsed/refractory FLT3-ITD+ AML. We report results from the global, randomized, double-blind, phase 3 QuANTUM-First trial (NCT02668653), evaluating quizartinib plus standard induction and post-remission consolidation (including allogeneic hematopoietic cell transplant [allo-HCT] in first complete remission [CR1]) followed by single-agent continuation (up to 3 years) vs placebo plus chemotherapy in newly diagnosed FLT3-ITD+ AML. Patients 18-75 years with newly diagnosed FLT3-ITD+ AML received induction with cytarabine 100 or 200 mg/m2/day (days 1-7) and daunorubicin 60 mg/m2/day or idarubicin 12 mg/m2/day (days 1-3). Patients were randomized to quizartinib (40 mg/day [days 8-21]) or placebo; stratified by region, age, and white blood cell count at diagnosis. Patients with CR or CR with incomplete hematologic recovery (CRi) received up to 4 cycles of high-dose cytarabine plus quizartinib (40 mg/day) or placebo and/or allo-HCT followed by continuation with quizartinib (30-60 mg/day) or placebo (up to 3 years). The primary endpoint was OS. Between September 2016 and August 2019, 539 patients were randomized (quizartinib [n=268], placebo [n=271]). Median age was 56 years (range, 20-75 years). As of August 2021 (median follow-up, 39.2 months), 58 patients remained on continuation. Median OS was significantly longer with quizartinib vs placebo (31.9 vs 15.1 months; HR, 0.776; 95% CI, 0.615-0.979; 2-sided P=.0324). CR+CRi rates were 71.6% and 64.9%, respectively. Allo-HCT in CR1 was performed in 157 patients (quizartinib, 31%; placebo, 27%). When censored for allo-HCT, OS trended longer with quizartinib vs placebo (HR, 0.752; 95% CI, 0.562-1.008; 2-sided P=.055). Relapse-free survival was longer with quizartinib vs placebo (HR, 0.733; 95% CI, 0.554-0.969). Grade ≥3 adverse events (AEs) were similar across arms. Discontinuations due to AEs occurred in 20.4% (quizartinib) and 8.6% (placebo). Fifty-six patients died from treatment-emergent AEs (quizartinib, 11.3%; placebo, 9.7%); mostly infections. Grade 3/4 electrocardiographic QT prolongation occurred in 3.0% (quizartinib) and 1.1% (placebo). Quizartinib plus standard therapy, followed by continuation, including after allo-HCT, for up to 3 years was tolerable with statistically significant and clinically meaningful OS improvements in adults ≤75 years with newly diagnosed FLT3-ITD+ AML.

AML-109 Preliminary Results of MT-401 in Post-Transplant Measurable Residual Disease Positive (MRD+) Acute Myeloid Leukemia (AML) Patients

Measurable residual disease (MRD) testing has become more prevalent in AML. MRD positivity is associated with increased relapse risk and shorter survival in AML and, currently, there are no approved therapies for these patients. To determine whether MRD+ patients post-allogeneic transplant can be treated safely and effectively with multi-tumor-associated antigen (mTAA)-specific T cells (MT-401/zedenoleucel) Design: A multicenter Phase 2 study (ARTEMIS) evaluating the safety and efficacy of MT-401 in patients with AML post-transplant is ongoing and includes patients with MRD+ disease. Three MRD+ AML patients were enrolled and treated in the ARTEMIS study. MRD testing was done by flow cytometry or molecular testing (e.g., RT-PCR). Patients received up to 3 consecutive infusions of MT-401 as a monotherapy (50-200×106 cells every 2 weeks) at weeks 0, 2, and 4 during the Intervention Period, and entered Follow-up starting at Week 8 for up to 5 years. The primary endpoint includes various safety measurements and incidence of dose-limiting toxicities (DLTs). The secondary endpoint was efficacy evaluations using ELN recommendations for standard AML response criteria. No DLTs were observed. The MRD+ patient had a variety of genetic mutations/abnormalities. One patient with NPM1 mutation converted from MRD+ to MRD- at week 8 evaluation. Another patient with RUNX1 genetic abnormality showed a decrease in MRD by PCR from a starting baseline of 0.8093% to resolution via peripheral blood at approximately 32 weeks post-treatment with MT-401. T cell receptor (TCR) analysis identified 3,117 antigen-specific clones (881 Survivin, 783 NY-ESO-1, 750 PRAME, 709 WT1). Immune monitoring of this patient showed T cell specificity not only for the targeted antigens but also for non-targeted antigens over time, thereby demonstrating epitope spreading. Interestingly, the tumor antigen composition by RNASeq identified an antigen expression profile that changes over time and inversely correlates with the presence of antigen-specific T cells, demonstrating the interplay of tumor cell immunogenicity and antigen-specific T cells. The lead-in portion of the ARTEMIS results showed that administration of MT-401 converted MRD+ patients to MRD-indicating that early intervention with MT-401 administration at MRD+ stage in post-transplant AML can be beneficial.