Form Of Acute Myeloid Leukemia Research Articles

Acute megakaryoblastic leukemia (AML7) about a case

Acute myeloid megakaryoblastic leukemia (AML7) is a rare and severe form of acute myeloid leukemia. Its diagnosis is based essentially on cytometric and cytogenetic data, given the complexity of morphological diagnosis. Myelofibrosis and osteolytic lesions may be a particular feature of AML7. Treatment with chemotherapy is considered, but the outcome is often unfavorable. The gold standard of treatment is bone marrow transplantation. Our article reports the case of an infant diagnosed with acute megakaryoblastic leukemia initially revealed by a bone marrow failure syndrome and aggravated by a cerebro-orbital extension with osteolytic lesions of the orbital cavity, treated with chemotherapy with a good clinical-biological evolution.

A Carbon 21 Steroidal Glycoside with Pregnane Skeleton from Cynanchum atratum Bunge Promotes Megakaryocytic and Erythroid Differentiation in Erythroleukemia HEL Cells through Regulating Platelet-Derived Growth Factor Receptor Beta and JAK2/STAT3 Pathway.

Erythroleukemia is a rare form of acute myeloid leukemia (AML). Its molecular pathogenesis remains vague, and this disease has no specific therapeutic treatments. Previously, our group isolated a series of Carbon 21 (C-21) steroidal glycosides with pregnane skeleton from the root of Cynanchum atratum Bunge. Among them, we found that a compound, named BW18, can induce S-phase cell cycle arrest and apoptosis via the mitogen-activated protein kinase (MAPK) pathway in human chronic myeloid leukemia K562 cells. However, its anti-tumor activity against erythroleukemia remains largely unknown. In this study, we aimed to investigate the anti-erythroleukemia activity of BW18 and the underlying molecular mechanisms. Our results demonstrated that BW18 exhibited a good anti-erythroleukemia activity in the human erythroleukemia cell line HEL and an in vivo xenograft mouse model. In addition, BW18 induced cell cycle arrest at the G2/M phase and promoted megakaryocytic and erythroid differentiation in HEL cells. Furthermore, RNA sequencing (RNA-seq) and rescue assay demonstrated that overexpression of platelet-derived growth factor receptor beta (PDGFRB) reversed BW18-induced megakaryocytic differentiation in HEL cells, but not erythroid differentiation. In addition, the network pharmacology analysis, the molecular docking and cellular thermal shift assay (CETSA) revealed that BW18 could inactivate Janus tyrosine kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway, which might mediate BW18-induced erythroid differentiation. Taken together, our findings elucidated a novel role of PDGFRB in regulating erythroleukemia differentiation and highlighted BW18 as an attractive lead compound for erythroleukemia treatment.

Acute Lymphoblastic Leukemia to Acute Promyelocytic Leukemia: phenotype switch at relapse. A Case Report

Lineage switch in acute leukemias is considered a rare event, especially in what concerns adult disease compared to pediatric patients. Differential diagnosis between biphenotypic/bilineal leukemias, de novo leukemias, therapy-related and phenotype switch variants, stands as an intricate challenge. Complementary laboratory studies and complex laboratory assays (conventional karyotyping, FISH- fluorescence in situ hybridization, molecular biology, immunophenotyping, etc.) represent one of the crucial resources regarding the process of establishing an accurate diagnosis, thus having a firm ground for the most beneficial treatment choice. Aim: To emphasize the diagnostic and therapeutic particularities regarding the case of a female patient with acute lymphoblastic leukemia (ALL) who later developed a phenotype switch towards an intermediary form of acute myeloid leukemia (AML), sharing features of both AML with maturation and atypical acute promyelocytic leukemia (AML M2/M3/APL). Material and methods: We hereby present the case of a 66 year old female, diagnosed with B- ALL, who underwent both classical chemotherapy (adequate protocol related to age, genetic profile, commorbidites) and bispecific monoclonal antibody therapy at first early relapse (Blinatumomab), followed by phenotype switch at second early relapse towards atypical APL. Combined ATRA and anthracycline therapy followed, yet laboratory work-up shows refractory disease. Complications during treatment were comprised of severe DIC (disseminated intravascular coagulation) and infections- sepsis followed by exitus. Results and conclusions: This is the case of a phenotype switch in a patient with ALL, who underwent both classical chemotherapy and Blinatumomab, towards myeloid lineage-refractory APL atypical variant. Availability of flow cytometry and cytogenetics during morphology evaluation represents a key aspect in the management of such hematologic malignancies.

The Genomic and Transcriptomic Landscape of Myeloid Sarcoma and Associated Acute Myeloid Leukemia

Background: Myeloid sarcoma (MS) is a distinct form of acute myeloid leukemia (AML) found in ~10% of patients, in which mass-forming myeloid blasts proliferate in extramedullary sites. MS may present as an isolated lesion as the primary manifestation of disease, co-occur with AML, or, more commonly, arise in the setting of AML relapse. It is associated with poor outcomes, and treatment options are sparse. While increased knowledge of AML and its biology have improved the care and survival of AML patients, those with MS continue to have limited successful treatment options, as MS has typically not been included in AML genetic and genomic profiling efforts, and patients are excluded from clinical trials. Leukemia cutis (LC) is a form of extramedullary AML characterized by cutaneous involvement, often with an infiltrative and non-mass forming pattern of growth. Still, historically the literature has not made a clear distinction between MS and LC. Although recent small pilot studies have suggested differences in AML-associated mutations between medullary AML and MS, knowledge of the complete molecular and cellular composition of MS and comprehensive comparison with AML is lacking. While molecular characterization of the extramedullary disease site is encouraged in the updated clinical recommendations, no definitive guidance exists for changes in the management and care of AML patients with MS, given the substantial gap in our current knowledge and understanding of extramedullary AML. Methods: Here, we performed RNA-Seq (n=96), single-cell (sc) RNA-Seq (n=33), and mutation profiling using either whole exome sequencing (WES, n=6) or a 409-gene targeted sequencing panel (n=48) of MS or LC patient samples and bone marrow (BM) aspirates from AML patients with and without extramedullary disease to present the first comprehensive characterization of MS and LC. Results: Our WES data showed thatMS and LC evolve independently in the extramedullary site from concurrent medullary disease with distinct dominant clones, characterized by higher median tumor mutation burden (1.66 vs. 064 mutations per megabase, respectively; p=0.0059) and increased copy number aberrations, which both validated in an MS Tet2 HR mouse model. Notably, we observed location-specific molecular evolution in patients with multiple concurrent MS sites (Figure 1A) and patients with recurring MS in WES and targeted panel data. While targetable mutations (i.e., IDH1/2 mutations and FLT3-ITD/TKD) occur, the inter-site variability of molecular aberrations may explain the short-lived responses of AML patients with MS to IDH/FLT3 inhibitors. Transcriptional profile differences between MS and associated medullary disease included up-regulation of extracellular matrix organization and cell-substrate adhesion and down-regulation of inflammatory response pathways in both bulk and scRNA-seq data (Figure 1B). Further, involved BM malignant cells of MS patients compared to patients without MS up-regulated specific pathways, including interferon-gamma response observed in both bulk and scRNA-seq, as well as chromatin organization and splicing. We further revealed that MS patients without overt BM involvement show remodeling of the BM immune microenvironment, with increased fractions of hematopoietic stem and progenitor cells that down-regulate HLA class II genes. Comparing LC and MS in the skin revealed distinct differences in their expression profiles, including high expression of genes involved in cell migration and immune response in LC, suggesting LC and MS represent distinct disease entities. Lastly, we demonstrated high expression of BCL2 in extramedullary disease (p=0.0074) and presented initial evidence that treatment with BH3-mimetics may be beneficial for the treatment of isolated MS. Conclusions: Myeloid sarcoma shows distinct molecular evolution and microenvironmental composition compared to medullary disease, including location-specific evolution necessitating personalized management consideration. Leukemia cutis is distinct from MS tumorous lesions and should be considered a separate entity. High BCL2 expression in extramedullary disease might represent a promising targetable vulnerability in patients with isolated MS.

Podoplanin Expression Measured By Flow Cytometry Accurately Identifies Patients with Acute Promyelocytic Leukemia at Diagnosis

Acute promyelocytic leukemia (APL) is a hematological emergency associated with high early mortality rates, thus requiring prompt diagnosis and treatment initiation. Currently, diagnostic suspicion relies on morphological criteria, which can be further corroborated by immunophenotypic analyses, until results based on molecular techniques confirm the diagnosis. However, while both cytology and immunophenotypic analyses are very important in this setting, their accuracy can be influenced by inherent morphological variation of blasts, as well as by gating and other technical issues related to flow cytometry. In this context, novel parameters that could contribute to the segregation of APL from other forms of acute myeloid leukemia (AML) would be desirable. Recently, Lavallée et al demonstrated that the aberrant expression of podoplanin (PDPN), a protein that mediates platelet activation in inflammatory contexts, by leukemic cells is as a relevant mediator of the coagulopathy of APL, as well potential biomarker of this condition based on the analysis of primary cells from patients with AML. The aim of this study was to prospectively and independently validate whether the surface expression of PDPN by leukemic promyelocytes measured by flow cytometry can contribute to the differential diagnosis between APL and other forms of AML. We also explored the association of PDPN expression with clinical and laboratory characteristics of these patients. The study population consisted of all consecutive patients diagnosed with APL by cytological, immunophenotypical and molecular techniques in our adult academic hematology hospital from 2020 to 2023. Exclusion criteria included the presence of hemodynamic instability or refusal to participate. The expression of PDPN was measured by flow cytometry using BD FACSCanto II™ cytometer. Analysis was performed in accordance with EuroFlow protocols, with a panel that includes CD45, CD33, CD34, MPO, CD117, CD13, HLA-DR, as well as an antibody against PDPN (PE anti-human Podoplanin Antibody - Catalog# 337004 - BioLegend). Lymphocytes were used as internal negative control for PDPN expression. Data were analyzed with Infinicyt™ software and reported both as the % of PDPN + cells or according to mean fluorescence intensity (MFI). Patients with other forms of AML diagnosed during the same period and matched for age, gender and peripheral blast count were used to compare the diagnostic accuracy of PDPN measurement. Clinical and laboratory data were obtained from the medical records. The study was approved by the Institutional Review Board and all participants provided written informed consent. In total, 19 consecutive patients with confirmed diagnosis of APL were compared with 18 patients with non-APL AML. Clinical and laboratory data from the study population are shown in table 1. The expression of PDPN in leukemic blasts was significantly higher in APL compared to non-APL AML, considering either the percentage of PDPN + cells or the MFI (table 1; figure 1a-b). When considered as a categorical variable, the expression of PDPN was strongly associated with the diagnosis of APL (Chi-square test= 19.169; P<0.001). PDPN expression was inversely correlated with platelet count (R S= -0.47; P=0.003) and fibrinogen (R S= -0.51; P=0.001), but not with other laboratory parameters such as PT and aPTT. No association of PDPN expression with the available clinical variables was observed, although it should be noted that our study is underpowered to confirm these findings. We next estimated the diagnostic accuracy of PDPN expression by flow cytometry by ROC analysis. As shown in figures 1c-d, the area under the ROC curve was higher than 0.9 for both analytical strategies. In conclusion, our prospective study demonstrated that measurement of the expression of PDPN by flow cytometry at diagnosis is capable to accurately segregate APL from other forms of AML. Additional studies are warranted to explore the clinical association of PDPN expression in larger and independent cohorts.

Hypercalcemia associated with interaction between all trans retinoic acid and fluconazole in an acute promyelocytic leukemia and acquired hypoparathyroidism case.

Acute promyelocytic leukemia (APL) is the most malignant form of acute myeloid leukemia (AML) with short survival without treatment. All trans retinoic acid (ATRA) is a vitamin A metabolite and plays an important role in the treatment of APL. Hypercalcemia is a rare side effect of ATRA. A 67-year-old female patient was investigated due to widespread bruising and pancytopenia. The patient was diagnosed with APL and remission was achieved by administering idarubicin together with ATRA in the induction treatment. The patient has hypocalcemia due to acquired hypoparathyroidism, and it was observed that the calcium level increased with the initiation of fluconazole 200 mg/day for antifungal prophylaxis together with ATRA in the consolidation treatment. It was observed that the calcium value reached 13 mg/dL by increasing the fluconazole to 400 mg/day treatment dose due to oral mucositis. The development of hypercalcemia has been reported in previous case reports when ATRA is used together with voriconazole, fosfluconazole, itraconazole, and posaconazole, which inhibit cytochrome P450 enzymes. In this case, it is the first in the literature that a patient with hypocalcemia due to acquired hypoparathyroidism developed hypercalcemia after fluconazole and ATRA were used together. Since hypercalcemia may develop while azole drugs are administered during ATRA treatment, it is important to monitor calcium levels to prevent complications of hypercalcemia.

Downregulation of stromal syntenin sustains AMLdevelopment.

The crosstalk between cancer and stromal cells plays a critical role in tumor progression. Syntenin is a small scaffold protein involved in the regulation of intercellular communication that is emerging as a target for cancer therapy. Here, we show that certain aggressive forms of acute myeloid leukemia (AML) reduce the expression of syntenin in bone marrow stromal cells (BMSC). Stromal syntenin deficiency, in turn, generates a pro-tumoral microenvironment. From serial transplantations in mice and co-culture experiments, we conclude that syntenin-deficient BMSC stimulate AML aggressiveness by promoting AML cell survival and protein synthesis. This pro-tumoral activity is supported by increased expression of endoglin, a classical marker of BMSC, which in trans stimulates AML translational activity. In short, our study reveals a vicious signaling loop potentially at the heart of AML-stroma crosstalk and unsuspected tumor-suppressive effects of syntenin that need to beconsidered during systemic targeting of syntenin in cancer therapy.

A rare case of acute basophilic leukemia

Acute basophilic leukemia is an extremely rare form of acute myeloid leukemia and diagnosis is based on morphological, cytochemical and ultrastructural analyses of basophils and blastic cells. It is usually characterized by a very rapid clinical course, symptoms of hyperhistaminemia and resistance to therapy. A 73 year-old male presented with itching, thrombocytopenia and leukocytosis. There was no hepatosplenomegaly. In the flow cytometry evaluation of peripheral blood, CD123 positive myeloid blasts were detected at the rate of 60%. Also, CD13 and CD33 were positive, while CD34, CD117, HLA DR and myeloperoxidase (MPO) were negative. These findings were considered as consistent with ABL. Following, the diagnosis of ABL was confirmed by bone marrow evaluation including cytomorphological and immunohistochemical studies. On the other hand, cytogenetical analyses of bone marrow showed the presence of the Philadelphia chromosome.The patient received one cycle of a chemotherapy regimen including cytarabine and idarubicin with imatinib support, but he died on the 10th hospital day. Considering that ABL behaves in a very malignant fashion, and a more aggressive treatment approach is necessary for patients with this specific subtype of AML.

Demographic factors affecting patients with AML with myelodysplasia-related changes.

e19027 Background: Acute myeloid leukemia with myelodysplasia-related changes (AML-MRC) is a rare and aggressive form of acute myeloid leukemia (AML) that makes up about 25-34% of its annual cases. The subtype includes a varied group of tumors with clinical, genetic, and morphological characteristics that confer poor outcomes. AML-MRC is commonly treated with intensive chemotherapy, but the median survival is only 9-12 months. Given the poor outcomes, gaining a better understanding of epidemiologic factors could reveal opportunities for earlier diagnosis and improved treatment planning. Demographic characteristics of AML-MRC patients were identified and examined using the National Cancer Database (NCDB). Methods: Individuals with a confirmed diagnosis of AML-MRC (N = 14,536) were identified using the NCDB for a retrospective analysis from 2004 to 2020. Patient sex, age at diagnosis, race, household income, insurance status, urban/rural residence, and Charlson-Deyo score were examined by descriptive statistics, and diagnostic trends by year were identified and calculated with regression analysis. Results: A total of 14,536 patients diagnosed with AML-MRC between 2004 – 2020 were identified in the NCDB. Within this timeframe, there was an increasing incidence of diagnosis per year (R2 = 0.9). AML-MRC was more common in males (61.1%) than females (38.9%), and the average age at diagnosis was 68.9 years (SD = 13.0, range = 0 – 90 years). The majority of patients (87.3%) were White. Patients were most likely to be a part of the highest median household income quartile (33.6%) and least likely to be a part of the bottom quartile (13.2%). A higher percentage of patients were insured by Medicare (63.0%) compared to private insurance (25.3%). Most patients lived in counties in metro areas of 1 million population or more (51.4%). Most individuals had Charlson-Deyo comorbidity scores of zero (68.1%). The most common type of treatment facility was academic/research programs (55.1%) followed by comprehensive community cancer programs (23.3%). Conclusions: AML-MRC is a common subtype of AML with a poor prognosis. Previous studies have found that most individuals with AML-MRC are older-aged, White males. Our study corroborated this finding and examined additional demographic factors. We found that these patients were more likely to hold insurance through Medicare, likely reflecting the older average age at diagnosis. Additionally, patients were most commonly in the highest household income quartile and living in a metropolitan area with the highest population density. This could suggest that better access to healthcare improves rates of diagnosis of AML-MRC. Especially given that more people are being diagnosed each year, further research should work to understand the effects of these, and other, socioeconomic factors on the diagnosis, treatment, and prognosis of patients with AML-MRC.

Acute Promyelocytic Leukemia Treatment Masking Hepatic Tuberculosis: A Management Dilemma

Acute promyelocytic leukemia is a form of acute myeloid leukemia (AML) that is characterized by presence of a promyelocytic leukemia-retinoic acid receptor alpha fusion. In most patients, this fusion is detected on conventional karyotype as the t(15;17)(q24.1;q21.2) translocation, but some patients have cryptic translocations with a normal karyotype. Historically, AML is associated with a poor prognosis. Treatment with all-trans retinoic acid and arsenic trioxide assures long-term survival in the majority of patients. This treatment is generally well-tolerated but may cause hepatotoxicity. This is usually identified by transaminitis but resolves after temporary cessation of treatment. Our patient's hepatotoxicity did not resolve following all-trans retinoic acid and arsenic trioxide cessation which posed a diagnostic dilemma. This prompted exploration of other possible causes of hepatotoxicity. An eventual liver biopsy identified acid-fast bacilli, confirming a diagnosis of hepatic tuberculosis. A broad differential diagnosis is imperative when investigating abnormalities in liver function, especially in chemotherapy patients when treatment cessation may cause cancer progression.

Erythroleukemia: Classification.

Acute erythroid leukemia (AEL) is a rare (2%-5%) form of acute myeloid leukemia (AML). Molecular alterations found in AEL resemble those of other AMLs. We report a classification of AELs in three major classes, with different prognosis and some specific features such as a tendency to mutual exclusion of mutations in epigenetic regulators and signaling genes.

A rare form of acute myeloid leukemia

A rare form of acute myeloid leukemia

A Novel Chalcone Derivative Regulates the Expression and Phosphorylation of ERK1/2 by Inhibiting Fli-1 Promoter Activity for Preventing the Malignant Progression of Erythroleukemia

Acute erythroleukemia is a rare form of acute myeloid leukemia recognized by its distinct phenotypic attribute of erythroblasts proliferation. In this study, in vitro experiments showed that a newly synthesized chalcone (ZH-254) inhibited cell proliferation, caused apoptosis, arrested the cell cycle in the G1 phase, and downregulated Fli-1 expression by inhibiting Fli-1 promoter activity. In vivo experiments showed that ZH-254 could effectively alleviate splenomegaly and prolong the survival of erythroleukemia mice. RT-PCR and Western blot analysis showed that ZH-254 could regulate the expression of Fli-1 target genes and G1-phase-related cell cycle proteins, including Rb, Bcl-2, Bax, ERK1/2, Gata-1, P110, SHIP-1, p-ERK1, CDK4, C-myc, Cyclin D1, Smad-3, GSK-3, and p21. Among them, the compound most significantly regulated the expression and phosphorylation of ERK1, the target gene of Fli-1 involved in regulating cell proliferation and apoptosis. Thus, ZH-254 restricts the malignancy of erythroleukemia by causing the inactivation of Fli-1 expression via suppressing its promoter activity, further regulating the expression and phosphorylation of ERK1- and G1-phase-related genes. These results reveal the critical role of Fli-1 in the growth and survival of various hematological malignancies and point to chalcone derivatives as lead compounds for the development of anti-Fli-1 drugs for the treatment of erythroleukemia with overexpression of Fli-1.

Consolidation of High-Risk Acute Promyelocytic Leukemia (APL) in Community Infusion Centers: A Single Center Experience

Intro: APL is one of the most curable forms of acute myeloid leukemia. Whereas all-trans retinoid acid (ATRA) and arsenic trioxide (ATO) are the standard of care for treatment of low/intermediate risk APL (APL0406 trial, LoCoco et al. NEJM 2013), the optimal treatment of high-risk APL remains unclear. In our institution, we use the APML4 regimen (Iland et al. Blood 2012) to treat high-risk APL patients, which uses idarubicin+ATRA followed by ATRA+ATO for induction. Then, there are two consolidation cycles of ATRA+ATO, the first of which is 28 consecutive days of treatment. Consolidation is followed by two years of ATRA + 6-mercaptopurine + methotrexate maintenance (PETHEMA LPA99, Sanz et al. Blood 2003). In the APML4 trial, the outcomes were excellent for all risk groups, particularly for high-risk patients. However, this regimen cannot be administered in community infusion centers (CIC) that are not open on weekends. In addition, the expertise required to give maintenance therapy makes it challenging to administer outside of an academic medical center. Here, we examine outcomes of high-risk APL patients who received induction at our institution to document the types of consolidation regimens they received and their outcomes. In particular, we focus on the outcomes of high-risk APL patients who received induction with idarubicin+ATRA+ATO (as per APML4) at our institution, and then for practical considerations received consolidation with ATRA+ATO (as per APL0406) in the community. Methods: We performed a retrospective analysis of all patients between the ages of 18-89 who were diagnosed with high-risk APL at the Knight Cancer Institute between January 2009 and July 2022. We identified 19 patients with the diagnosis of high-risk APL. One patient died soon after diagnosis due to intracranial bleed (excluded from final analysis). The remainder completed induction and consolidation. Result: Table 1 shows the main clinical features at diagnosis. The most common complications during induction were differentiation syndrome and coagulopathy. All 18 patients achieved complete morphologic remission (CR) at the end of induction but still had detectable PML-RARA by PCR. 4/18 (22%) patients received the APL2000 regimen (Adès et al. JCO 2006), and 1/18 (6%) patient received the PETHEMA regimen (Sanz et al. Blood 2003). All 5 patients were hospitalized for neutropenic fever, which required significant dose reduction and treatment delay. One patient (6%) received induction with ATRA+ATO+gemtuzumab ozogamicin followed by consolidation with ATRA+ATO (Estey et al. Blood 2006). The majority of patients (12/18, 67%) received induction as per APML4 protocol (Table 2). The APL0406 regimen was administered as consolidation in 11/18 (61%) patients; and all patients except one received consolidation at local CIC. Common side effects were headache and nausea, leading to dose reduction in five patients. One patient continued with APML4 consolidation. Molecular CR was confirmed in all patients at the end of consolidation treatment. 13/18 (72%) patients did not receive maintenance therapy after consolidation. All patients were subsequently followed by RT-PCR for PML/RARA for molecular relapse. At the last follow up, all 18 patients remained to be in molecular CR with no relapsed disease. Discussion: The benefit of ATRA+ATO for low/intermediate risk APL patients is well established, but the optimal management of high-risk APL remains a challenge, especially when consolidation is given in CIC. Our series of patients provides real world evidence that ATRA+ATO consolidation as per APL0406 regimen is effective in high-risk APL patients after induction with idarubicin+ATRA+ATO as per the APML4 regimen. Although the numbers are limited, all patients remain in molecular remission as measured from the end of consolidation, similar to the excellent outcomes of low/intermediate risk APL patients. Consolidation was well tolerated, with minimal treatment delays, suggesting that a standard consolidation regimen for low-, intermediate-, and high-risk APL patients can be successfully administered in the community (most centers typically treat 1-2 patients per year with APL). The majority of patients did not receive maintenance therapy but remained in remission, providing further evidence that maintenance therapy is unnecessary with sufficient ATRA+ATO consolidation, and simplifying therapy (Shah et al. Blood 2018). Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Aberrant EVI1 splicing contributes to EVI1-rearranged leukemia

AbstractDetailed genomic and epigenomic analyses of MECOM (the MDS1 and EVI1 complex locus) have revealed that inversion or translocation of chromosome 3 drives inv(3)/t(3;3) myeloid leukemias via structural rearrangement of an enhancer that upregulates transcription of EVI1. Here, we identify a novel, previously unannotated oncogenic RNA-splicing derived isoform of EVI1 that is frequently present in inv(3)/t(3;3) acute myeloid leukemia (AML) and directly contributes to leukemic transformation. This EVI1 isoform is generated by oncogenic mutations in the core RNA splicing factor SF3B1, which is mutated in >30% of inv(3)/t(3;3) myeloid neoplasm patients and thereby represents the single most commonly cooccurring genomic alteration in inv(3)/t(3;3) patients. SF3B1 mutations are statistically uniquely enriched in inv(3)/t(3;3) myeloid neoplasm patients and patient-derived cell lines compared with other forms of AML and promote mis-splicing of EVI1 generating an in-frame insertion of 6 amino acids at the 3′ end of the second zinc finger domain of EVI1. Expression of this EVI1 splice variant enhanced the self-renewal of hematopoietic stem cells, and introduction of mutant SF3B1 in mice bearing the humanized inv(3)(q21q26) allele resulted in generation of this novel EVI1 isoform in mice and hastened leukemogenesis in vivo. The mutant SF3B1 spliceosome depends upon an exonic splicing enhancer within EVI1 exon 13 to promote usage of a cryptic branch point and aberrant 3′ splice site within intron 12 resulting in the generation of this isoform. These data provide a mechanistic basis for the frequent cooccurrence of SF3B1 mutations as well as new insights into the pathogenesis of myeloid leukemias harboring inv(3)/t(3;3).

Abstract 2759: CAR-T engager proteins for the treatment of B cell cancers and acute myeloid leukemia

Abstract Chimeric Antigen Receptor T cells targeting malignancies expressing CD19 (CAR19) have been widely successful, with products approved to treat B cell lymphomas (NHL) and B cell leukemia. A major limitation of CAR19 therapy is the steep relapse rate within 6 months of treatment, often due to the loss or diminution of tumor cell CD19 expression. ALETA-001 is a CAR-T Engager protein that contains the CD19 extracellular domain (ECD), an anti-CD20 VHH, and an anti-albumin VHH for half-life extension. When combined with CAR19 T cells, ALETA-001 triggers cytotoxicity through CD19 bound to CD20, thus increasing total target antigen density and preventing relapse due to loss of CD19 expression. ALETA-001 will enter Cancer Research UK-sponsored Phase 1/2 clinical trials in CAR19-treated NHL patients next year. We made novel variants of ALETA-001 including CAR-T Engagers with alternative anti-CD20 modules and CAR-T Engagers that target both CD20 and a second B cell cancer antigen. Further, we extended our technology to target Acute Myeloid Leukemia (AML), using novel antigen binding domains and immunomodulatory functional domains. Site-directed mutagenesis was used to make ALETA-001 variants. We mutated the complementary determining regions, CDR2 and/or CDR3, within the anti-CD20 VHH. The novel CAR-T Engager Proteins were evaluated for binding to CD19-negative/CD20-positive lymphoma cells and for cytotoxicity against those cells in the presence of CAR19 T cells. Further, CAR-T Engagers were made that contained novel CD20 binding domains and binding domains to other B cell antigens. In addition, we created a novel CAR-T engager that contains the CD19 ECD and binding domains to two AML antigens, plus an immunomodulatory domain. This novel CAR-T engager is designed to treat CD19-positive Mixed Phenotype Leukemia (MPL) and other forms of AML. We evaluated the series of CDR mutations in the ALETA-001 anti-CD20 VHH. A spectrum of activities was observed, with mutations that retained full CD20 binding and cytotoxic activity to those with partial or no activity. A series of constructs with distinct anti-CD20 binding domains were also evaluated, with some found to be equipotent with ALETA-001. Constructs binding both CD20 and a second B cell tumor antigen were prepared and assayed and found to have potent activity against either or both antigens. Finally, CAR-T Engagers directed to AML antigens were able to bind each antigen individually, and mediated potent toxicity with the addition of CAR19 T cells. We conclude that the CAR-T Engager platform offers robust and modular anti-tumor functionality, featuring potent multi-antigen targeting for diverse indications. Finally, we can incorporate other useful immunomodulatory activities into Engager proteins. Citation Format: Paul D. Rennert, Lihe Su, Lan Wu, Roy R. Lobb, Christine Ambrose. CAR-T engager proteins for the treatment of B cell cancers and acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2759.

All-Trans-Retinoic Acid Combined With Valproic Acid Can Promote Differentiation in Myeloid Leukemia Cells by an Autophagy Dependent Mechanism.

Acute myeloid leukemia (AML) is an aggressive blood cancer with an overall survival of 30%. One form of AML, acute promyelocytic leukemia (APL) has become more than 90% curable with differentiation therapy, consisting of all-trans-retinoic acid (ATRA) and arsenic trioxide (ATO). Application of differentiation therapy to other AML subtypes would be a major treatment advance. Recent studies have indicated that autophagy plays a key role in the differentiation of ATRA-responsive APL cells. In this study, we have investigated whether differentiation could be enhanced in ATRA resistant cells by promoting autophagy induction with valproic acid (VPA). ATRA sensitive (NB4) and resistant leukemia cells (NB4R and THP-1) were co-treated with ATRA and valproic acid, followed by assessment of autophagy and differentiation. The combination of VPA and ATRA induced autophagic flux and promoted differentiation in ATRA-sensitive and -resistant cell lines. shRNA knockdown of ATG7 and TFEB autophagy regulators impaired both autophagy and differentiation, demonstrating the importance of autophagy in the combination treatment. These data suggest that ATRA combined with valproic acid can promote differentiation in myeloid leukemia cells by mechanism involving autophagy.

The evaluation of risk factors leading to early deaths in patients with acute promyelocytic leukemia: a retrospective study.

Acute promyelocytic leukemia (APL) differs from other forms of acute myeloid leukemia (AML), including coagulopathy, hemorrhage, disseminated intravascular coagulation (DIC), and treatment success with all-trans retinoic acid (ATRA). Despite ATRA, early deaths (ED) are still common in APL. Here, we evaluated factors associated with ED and applicability of scoring systems used to diagnose DIC. Ninety-one APL patients (55 females, 36 males, and median age 40 years) were included. ED was defined as deaths attributable to any cause between day of diagnosis and following 30th day. DIC was assessed based on DIC scoring system released by the International Society of Thrombosis and Hemostasis (ISTH) and Chinese Diagnostic Scoring System (CDSS). Patients' median follow-up time was 49.2 months, and ED developed in 14 (15.4% of) cases. Patients succumbing to ED had higher levels of the Eastern Cooperative Oncology Group Performance Status (ECOG PS), lactate dehydrogenase (LDH), and ISTH DIC, and lower fibrinogen levels (p <0.05). In multivariate Cox regression analysis, age >55 and ECOG PS ≥2 rates were revealed to be associated with ED. Based on ISTH and CDSS scores, DIC was reported in 47.3 and 58.2% of the patients, respectively. Despite advances in APL, ED is still a major obstacle. Besides the prompt recognition and correction of coagulopathy, those at high ED risk are recommended to be detected rapidly. Implementation of local treatment plans and creating awareness should be achieved in hematological centers. Common utilization of ATRA and arsenic trioxide (ATO) may be beneficial to overcome ED and coagulopathy in APL patients.

Euro-BioImaging – Interdisciplinary research infrastructure bringing together communities and imaging facilities to support excellent research

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Infiltration of thyroid papillary cancer tissue with myeloid leukemic cells: a case report

BackgroundExtramedullary leukemia, also known as myeloid sarcoma, is a rare form of acute myeloid leukemia and often accompanies bone marrow involvement. Myeloid infiltration of the thyroid gland is extremely rare. Here, a unique case in which thyroid cancer tissue was infiltrated with myeloid cells is presented.Case presentationWe present a case of thyroid papillary cancer infiltrated by blastic cells and bilateral breast and axillary myeloid sarcoma in a 30-year-old Caucasian female patient with a history of osteosarcoma and MDS-RAEB2. The patient firstly received 6 cycles of chemotherapy for osteosarcoma, and allogeneic hematopoietic stem cell transplantation was performed after anthracycline-based chemotherapy due to MDS-RAEB2. The patient remained in remission on follow-up in terms of both osteosarcoma and MDS-RAEB2. Malignant features (Bethesda VI) were observed in the fine needle aspiration biopsy performed from a newly developed firm, fixed thyroid nodule approximately 4–5 cm in length in the left thyroid lobe. Because of the Bethesda VI thyroid nodule, the patient underwent total thyroidectomy. In the pathological evaluation, CD34-, CD117-, MPO-, and HLA-DR-positive blastic cells which infiltrated into follicular variant papillary thyroid carcinoma were detected. In the evaluation performed due to blastic cell infiltration, multiple lesions showing increased 18-fluorodeoxyglucose activity in bilateral breast and axillae were detected. Myeloid sarcoma was found as a result of tru-cut biopsy from these lesions. A fungal cystic lesion was detected in the frontal region of the patient who developed altered consciousness after the second cycle of treatment of myeloid sarcoma. During her follow-up in the intensive care unit, she died of cranial septic embolism and acute infarction.ConclusionsHere, we present a very interesting case that is the first. A staged approach to diagnosis with methods including immunohistochemical staining, radiological imaging methods, and cytogenetic and molecular analyses can help make the definitive diagnosis.