Onset Of Acute Myeloid Leukemia Research Articles

Increased Th17 and Treg levels in peripheral blood positively correlate with minimal residual disease in acute myeloid leukaemia.

Immune dysregulation plays a key role in acute myeloid leukemia (AML). We aimed to explore the correlation between T helper cell 17 (Th17) and the regulatory cells (Tregs) in the peripheral blood of patients with newly diagnosed (ND) AML and bone marrow blast cells, as well as minimal residual disease (MRD) before and after treatment. Changes in Th17 and Treg cells in the peripheral blood of 32 patients with ND AML were observed before and after induction chemotherapy with cytarabine for seven days and anthracycline for three days. The levels of inflammatory cytokines were measured using an enzyme-linked immunosorbent assay. Correlation analysis between bone marrow blast cells and Th17 and Treg cell frequencies was performed using the Pearson's correlation test. Frequencies of Th17 and Treg cells and MRD were assessed using flow cytometry. IL-6, IL-10, IL-17A, and GM-CSF levels gradually increased in patients with ND AML and CR and NR patients. The percentages of Th17 and Treg cells positively correlated with those of blast cells. In addition, the frequencies of Th17 and Treg cells in MRD-positive patients were higher than those in MRD-negative patients at the initial induction and after three months of chemotherapy. The frequencies of Tregs and Th17 cells positively correlated with MRD onset. Increased Th17 and Treg cell levels were positively correlated with onset of AML, poor remission, and MRD.

The ISTH DIC-score predicts early mortality in patients with non-promyelocitic acute myeloid leukemia

Coagulation disorders frequently complicate the clinical course of acute myeloid leukemia (AML) patients. This study examined the frequency and prognostic significance, with regards of early mortality, of the presence of overt disseminated intravascular coagulation (DIC) at AML diagnosis and its correlation with clinical and biological characteristics.A retrospective analysis of 351 newly diagnosed non-promyelocytic AML patients was conducted, utilizing the 2018 ISTH DIC-Score criteria to evaluate the presence of overt DIC at AML onset. The study cohort had a median age of 65 years with a predominance of male gender (59 %). Overt DIC was present in 21 % of cases and was associated with advanced age, comorbidities, poor performance status, hyperleukocytosis, LDH levels, NPM1 mutations, expression of CD33 and CD4, and lack of expression of CD34. With a median follow-up of 72 months (3–147 months), the 6-year overall survival (OS) was 17.4 %, with patients having overt DIC showing significantly poorer outcomes (7.2 % compared to 20.3 % of those without DIC, p < 0.001). Patients with overt DIC showed markedly high early mortality rates at 30 (42.5 % vs 8 %), 60 (49.3 % vs 16.9 %), and 120 days (64.4 % vs 25.6 %) from disease onset. In multivariate analysis overt DIC retained its independent prognostic value for early mortality.In conclusion, the prevalence and clinical relevance of DIC in non-promyelocytic AML is not negligible, underlining its potential as an unfavorable prognostic marker. In newly diagnosed patients with AML, early recognition and measure to counteract coagulation disturbances might help mitigate the elevated mortality risk associated with DIC.

Ocular manifestations of acute myeloid leukemia during induction phase of chemotherapy: A case series

The aim of this series is to report diverse ocular features noted in patients during the induction phase in acute myeloid leukemia (AML). Three female patients undergoing induction therapy for AML reporting for various ocular complaints were examined. Detailed history, including the onset of AML and chemotherapy and any past ocular or systemic ailment, was taken. Best-corrected visual acuity, anterior segment, and posterior segment examinations were performed. Intraocular pressure was measured, and color vision was tested. Routine hematological, and biochemical investigations and imaging were done when required. Patients with abnormal parameters were referred to an oncologist for management. A varied spectrum of ocular manifestations was noted in these three female patients, including preseptal cellulitis, orbital cellulitis, and subhyaloid hemorrhages. Immediate treatment was given to patients with vision-threatening conditions in consultation with an oncologist and otorhinolaryngologist, and patients responded well to the treatment. Owing to its cytoreductive action, AML induction therapy exposes the patient to various adverse events, causing significant ocular and systemic morbidity. All the diagnosed patients of AML are recommended to undergo a baseline ocular assessment.

Clinical features and management of germline CEBPA-mutated carriers

Familial acute myeloid leukemia (AML) pedigrees with germline CCAAT/enhancer-binding protein-α (CEBPA) mutation have been rarely reported due to insufficient knowledge of their clinical features. Here, we report two Chinese families with multiple AML cases carrying germline CEBPA mutations, one of which had 11 cases spanning four consecutive generations. Additionally, we collected clinical data of 57 AML patients from 22 families with germline CEBPA mutations, with 58.3% of them harboring double CEBPA mutations. The first mutation frequently occurred at the N-terminal of CEBP/α (78.6%), resulting in an exclusive expression of p30 of CEBPA (CEBPAp30). The second mutation was mostly found at the C-terminal of CEBP/α (CEBPAothers). Germline CEBPAp30 carriers had higher incidences of AML (80.36% vs. 42.86%) and earlier onset of AML (18 vs. 38.5 years old) compared to germline CEBPAothers carriers. Despite the high rates of relapse, most familial AML cases exhibited favorable overall survival (OS), with germline CEBPAp30 carriers having better survival outcomes (>25 years vs. 11 years for CEBPAothers carriers). Among the 27 healthy germline CEBPA-mutated carriers, we detected a pre-leukemia clone harboring a pathogenic IDH2 variant (R140Q)in one individual. These findings should aid in the genetic counseling and management of AML patients and healthy carriers with germline CEBPA mutations.

Drug Repurposing in a Zebrafish AML Model

Acute myeloid leukemia (AML) is a genetically heterogeneous hematological malignancy. AML arises from the stepwise accumulation of mutations in hematopoietic stem cells (HSCs). The t(9;11)(p22;q23) translocation is a translocation detected in about 5 to 7% of AML patients, which gives rise to the KMT2A/MLLT3 or MLL/AF9 ( MA9) fusion gene. Zebrafish (ZF) are well-established model organisms for investigating human hematopoiesis. Approximately 70% of human genes have corresponding orthologues in zebrafish, and over 80% of disease-related genes are conserved between the two species. Previously, we established a transgenic MA9 zebrafish AML model (Runx1+23:MA9), in which the AML was serially transplantable. Our investigations revealed that MA9 expression alters hematopoiesis in transgenic embryos before the onset of AML, leading to myeloid expansion. Whole-mount in situ hybridization (WISH) of embryos showed increased expression of granulocyte markers such as lyz 24 and 48 hours after fertilization (hpf). Based on these findings, we developed a Runx1+23:MA9/lyz-EGFP zebrafish leukemia line, in which lyz-expressing myeloid cells are EGFP positive. This model provides a phenotype-based platform to evaluate the therapeutic potential of any drug that can affect the myeloid cell population. We used this ZF model to investigate the therapeutic potential of nine drugs in MA9-driven AML. MA9 transgenic fish (Runx1+23:MA9) were crossed with lyz-EGFP fish to generate a MA9/lyz-EFGP line. In this line, myeloid cells can be visualized with a fluorescent microscope. The expression of lyz in the MA9/lyz-EFGP transgenic embryos was compared to that in the lyz-EGFP line. In our previous study on MA9 transgenic leukemic fish, RNA-seq analysis had revealed higher expression of genes acting in the EGFR, MEK, RAF, KRAS pathways and downregulation of genes in TP53. Based on these results, we selected nine drugs that are known to affect some of these pathways as well as drugs that are used to treat leukemia for screening in our model. Control and MA9/lyz-EFGP ZF 48 hpf embryos were treated with 10 µM of each drug. 24 hrs after drug treatment, the number of lyz expressing cells was analyzed by visualizing the embryos under a fluorescent microscope, and lyz transcript levels were measured by qPCR. Treatment with erlotinib, lapatinib, cytarabine, and gefitinib led to a visible reduction in the number of lyz-expressing cells in the MA9 transgenic embryos but not in the controls (Figure 1). In comparison, treatment with decitabine resulted in a decrease in the number of lyz expressing cells in both the MA9/lyz-EGFP and the control embryos. Treatment with doxorubicin, zoledronic and arsenic trioxide did not have an effect on the number of lyz-expressing population in the transgenic nor in the control embryos. These results were confirmed by qPCR, which showed that expression of lyz was reduced by 33% to 51% in the erlotinib, lapatinib, cytarabine, and gefitinib treated MA9 transgenic embryos (Figure 2). Treatment with these drugs did not significantly change the lyz expression in lyz-EFGP control embryos. Our results provide strong evidence for the successful establishment of a zebrafish model of KMT2A-rearranged AML that can be used for low to medium-throughput drug screening by employing the expression level of an endogenous gene (lyz) as a read-out. This model is a valuable tool to assess the drug's specificity in targeting oncogene-expressing cells and normal hematopoietic cells and, therefore, to identify novel therapeutic agents that can target the cancer cells with fewer side effects on normal hematopoietic cells.

Wilms' Tumor 1 Functions As a Tumor Suppressor to Suppress FLT3-STAT Signaling and Epigenetic Remodeling in Acute Myeloid Leukemia (CALGB 8461, 9665 and 20202; Alliance)

The Wilms' Tumor 1 ( WT1) gene is a transcription factor that is recurrently mutated or commonly overexpressed in several cancer types. In acute myeloid leukemia (AML), frequent overexpression of WT1 and poor patient outcomes associated with WT1 mutations highlight its importance in the disease; however, there are no tailored treatments for these patients. Furthermore, WT1's fundamental role as either an oncogene or tumor-suppressor remains unresolved. Here we examine the roles of both wild-type and mutant WT1 in AML through epigenetic and mechanistic studies. Using our findings we propose a personalized treatment strategy. WT1 mutations are enriched in the NPM1 mut subset of AML; thus we first focused our study on a cohort of 581 patients with de novo AML and NPM1 mutations enrolled on Alliance for Clinical Trials in Oncology studies. Transcriptomic analyses revealed that WT1 mut patientsphenocopied a distinct, aberrant gene expression signature associated with FLT3 internal tandem duplications (ITD), a mutation known to activate STAT signaling and associated with poor outcome in AML. We observed that WT1 expression levels were remarkably elevated in FLT3-ITD patients and were driven by STAT5A binding to the WT1 promoter. STAT5A binding and subsequent WT1 upregulation were blocked by small molecule FLT3 inhibitors. These findings linking FLT3 activity with WT1 expression raise two possibilities: either WT1 is an oncogene cooperating with the FLT3-STAT pathway, or WT1 naturally functions to suppress FLT3 signaling in a negative feedback loop subverted by WT1 mutations. As WT1 interacts with TET2 to facilitate epigenetic remodeling and DNA binding, we performed genome-wide DNA methylation analysis on WT1 mut AML patients and observed selective hypermethylation of WT1 binding motifs consistent with loss of function. We confirmed that WT1 mutations in AML commonly cause truncation of the C-terminal DNA binding domain in our patient cohort. Using co-immunoprecipitation, ChIP-sequencing and luciferase reporter assays, we found that mutant WT1 functions as a dominant-negative, inhibiting WT1 derived from the wild-type allele. Investigation of WT1 target genes by intersecting transcriptomic and DNA methylation profiles identified hypermethylation and downregulation of miR-193a, leading to a significant downregulation of miR-193a-3p in WT1 mut AML patients. Overexpression of wild-type, but not mutated, WT1 rescued miR-193a expression in AML cell lines. Enforced expression of miR-193a significantly delayed AML onset in vivo. In addition, overexpression of miR-193a in multiple human AML cell lines and primary patient samples impaired AML cell growth and colony-forming capacity while promoting monocytic differentiation, underscoring its role as potent tumor suppressor. Therapeutic modulation of miRNA levels in cancer patients has been limited by inefficient delivery and tissue enrichment. To overcome this, we tested a novel lipid-nanoparticle (LNP) formulation of miR-193a-3p (INT-1B3), currently being investigated in a phase I clinical trial (NCT04675996). Biweekly i.v. treatments of INT-1B3 in the immunocompetent Hoxa9/Meis1 (H9M)-transduced model system prevented AML formation, highlighting the potent anti-leukemic activity of this miRNA based therapeutic. Overexpression of miR-193a-loaded LNPs downregulated FLT3 expression and suppressed STAT signaling in primary AML samples. Finally, treatment of primary AML cells with FLT3 inhibitors revealed enhanced sensitivity of WT1 mut cells in the absence of FLT3-ITD, highlighting the role of wild-type FLT3 in WT1 mut cells. In summary, we uncovered a critical negative-feedback loop maintained by WT1 to suppress FLT3 activity. Loss-of-function WT1 mutations subvert the tumor suppressor function of WT1 via failure to maintain miR-193a expression, leading to increased FLT3 expression and STAT5 signaling, subsequently impairing differentiation, increasing proliferation and disease aggressiveness in AML (Figure 1). Our findings advocate for use of FLT3 inhibition and miR-193a supplementation for treatment of WT1 mut patients, a subgroup with poor outcomes and no targeted treatment options.

A comprehensive analysis of nucleotide excision repair characteristics defines a novel prognostic signature for acute myeloid leukemia.

Nucleotide excision repair (NER) has been associated with various types of malignant tumors. However, the precise roles of nucleotide excision repair-related genes (NERGs) in acute myeloid leukemia (AML) remain incompletely understood. Hence, this study aimed to develop a prognostic signature incorporating NERGs in AML, which could potentially predict patient outcomes. By querying the Genotype-Tissue Expression (GTEx), The Cancer Genome Atlas (TCGA), and Gene Expression Omnibus (GEO) databases, we acquired RNA-seq data and clinical information pertaining to AML. To identify differentially expressed NERGs (DE-NERGs), we employed the Wilcoxon rank-sum test. Based on the expression patterns of DE-NERGs with prognostic significance, patients were categorized into two subgroups. A prognostic signature was developed through univariate Cox regression and least absolute shrinkage and selection operator (LASSO) analyses to compare the differentially expressed genes (DEGs) between these two groups. Additionally, a nomogram was constructed using multivariate analysis. The biological pathways involved were elucidated through Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, gene set variation analysis (GSVA), and gene set enrichment analysis (GSEA). We developed a prognostic model based on an 11-gene signature. Furthermore, the risk score derived from this model was demonstrated to independently serve as a prognostic marker for patients diagnosed with AML. Our prognostic model, based on NERGs, was developed and validated to provide insights into the onset and progression of AML and establish a foundation for more effective treatment. Our findings not only contribute to clinical decision-making but also underscore the significance of nucleotide excision repair. Furthermore, they may pave the way for the development of targeted therapeutic strategies specifically focused on this process.

Biallelic disruption of DDX41 activity is associated with distinct genomic and immunophenotypic hallmarks in acute leukemia.

Inherited DDX41 mutations cause familial predisposition to hematologic malignancies including acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS), with the majority of DDX41 mutated MDS/AMLs described to date harboring germline DDX41 and co-occurring somatic DDX41 variants. DDX41-AMLs were shown to share distinguishing clinical features such as a late AML onset and an indolent disease associated with a favorable outcome. However, genotype-phenotype correlation in DDX41-MDS/AMLs remain poorly understood. Here, we studied the genetic profile, bone marrow morphology and immunophenotype of 51 patients with DDX41 mutations. We further assessed the functional impact of ten previously uncharacterized DDX41 variants of uncertain significance. Our results demonstrate that MDS/AML cases harboring two DDX41 variants share specific clinicopathologic hallmarks that are not seen in other patients with monoallelic DDX41 related hematologic malignancies. We further showed that the features seen in these individuals with two DDX41 variants were concordant with biallelic DDX41 disruption. Here, we expand on previous clinicopathologic findings on DDX41 mutated hematologic malignancies. Functional analyses conducted in this study unraveled previously uncharacterized DDX41 alleles and further illustrate the implication of biallelic disruption in the pathophysiology of this distinct AML entity.

A Question of Frame: The Role of the Bone Marrow Stromal Niche in Myeloid Malignancies.

Until a few years ago, the onset of acute myeloid leukemia (AML) was entirely ascribed to genetic lesions in hematopoietic stem cells. These mutations generate leukemic stem cells, which are known to be the main ones responsible for chemoresistance and relapse. However, in the last years, increasing evidence demonstrated that dynamic interplay between leukemic cells and bone marrow (BM) niche is of paramount relevance in the pathogenesis of myeloid malignancies, including AML. Specifically, BM stromal niche components, such as mesenchymal stromal cells (MSCs) and their osteoblastic cell derivatives, play a key role not only in supporting normal hematopoiesis but also in the manifestation and progression of myeloid malignancies. Here, we reviewed recent clinical and experimental findings about how genetic and functional alterations in MSCs and osteolineage progeny can contribute to leukemogenesis and how leukemic cells in turn generate a corrupted niche able to support myeloid neoplasms. Moreover, we discussed how the newest single-cell technologies may help dissect the interactions between BM stromal cells and malignant hematopoiesis. The deep comprehension of the tangled relationship between stroma and AML blasts and their modulation during disease progression may have a valuable impact on the development of new microenvironment-directed therapeutic strategies, potentially useful for a wide cohort of patients.

Acute Myeloid Leukemia Causes Serious and Partially Irreversible Changes in Secretomes of Bone Marrow Multipotent Mesenchymal Stromal Cells.

In patients with acute myeloid leukemia (AML), malignant cells modify the properties of multipotent mesenchymal stromal cells (MSCs), reducing their ability to maintain normal hematopoiesis. The aim of this work was to elucidate the role of MSCs in supporting leukemia cells and the restoration of normal hematopoiesis by analyzing ex vivo MSC secretomes at the onset of AML and in remission. The study included MSCs obtained from the bone marrow of 13 AML patients and 21 healthy donors. The analysis of proteins contained in the MSCs-conditioned medium demonstrated that secretomes of patient MSCs differed little between the onset of AML and remission; pronounced differences were observed between MSC secretomes of AML patients and healthy donors. The onset of AML was accompanied by a decrease in the secretion of proteins related to ossification, transport, and immune response. In remission, but not at the onset, secretion of proteins responsible for cell adhesion, immune response, and complement was reduced compared to donors. We conclude that AML causes crucial and, to a large extent, irreversible changes in the secretome of bone marrow MSCs ex vivo. In remission, functions of MSCs remain impaired despite the absence of tumor cells and the formation of benign hematopoietic cells.

Phytochemicals and bioactive compounds effective against acute myeloid leukemia: A systematic review.

This systematic review identified various bioactive compounds which have the potential to serve as novel drugs or leads against acute myeloid leukemia. Acute myeloid leukemia (AML) is a heterogeneous hematopoietic malignancy that arises from the dysregulation of cell differentiation, proliferation, and cell death. The risk factors associated with the onset of AML include long-term exposure to radiation and chemicals such as benzene, smoking, genetic disorders, blood disorders, advancement in age, and others. Although novel strategies to manage AML, including a refinement of the conventional chemotherapy regimens, hypomethylating agents, and molecular targeted drugs, have been developed in recent years, resistance and relapse remain the main clinical problems. In this study, three databases, PubMed/MEDLINE, ScienceDirect, and Google Scholar, were systematically searched to identify various bioactive compounds with antileukemic properties. A total of 518 articles were identified, out of which 59 were viewed as eligible for the current report. From the data extracted, over 60 bioactive compounds were identified and divided into five major groups: flavonoids, alkaloids, organosulfur compounds, terpenes, and terpenoids, and other known and emerging bioactive compounds. The mechanism of actions of the analyzed individual bioactive molecules differs remarkably and includes disrupting chromatin structure, upregulating the synthesis of certain DNA repair proteins, inducing cell cycle arrest and apoptosis, and inhibiting/regulating Hsp90 activities, DNA methyltransferase 1, and histone deacetylase 1.

Orbital impairment in the debut of oncohematological diseases in children

Tumor lesion of the orbit is found in the clinic of various oncohematological diseases in children. Untimely referral of such patients to an oncologist delays the timing of the necessary treatment, thereby worsening the prognosis of the disease. The literature review presents the features of clinical manifestations of orbital pathology in children at the onset of acute myeloid leukemia, myeloid sarcoma, lymphoma, Langerhans cell and non-Langerhans cell histiocytosis. The article presents our own observations and identifies factors that negatively affect the timely diagnosis of oncohematological diseases in this category of patients.

The histone acetyltransferase KAT6A is recruited to unmethylated CpG islands via a DNA binding winged helix domain.

The lysine acetyltransferase KAT6A (MOZ, MYST3) belongs to the MYST family of chromatin regulators, facilitating histone acetylation. Dysregulation of KAT6A has been implicated in developmental syndromes and the onset of acute myeloid leukemia (AML). Previous work suggests that KAT6A is recruited to its genomic targets by a combinatorial function of histone binding PHD fingers, transcription factors and chromatin binding interaction partners. Here, we demonstrate that a winged helix (WH) domain at the very N-terminus of KAT6A specifically interacts with unmethylated CpG motifs. This DNA binding function leads to the association of KAT6A with unmethylated CpG islands (CGIs) genome-wide. Mutation of the essential amino acids for DNA binding completely abrogates the enrichment of KAT6A at CGIs. In contrast, deletion of a second WH domain or the histone tail binding PHD fingers only subtly influences the binding of KAT6A to CGIs. Overexpression of a KAT6A WH1 mutant has a dominant negative effect on H3K9 histone acetylation, which is comparable to the effects upon overexpression of a KAT6A HAT domain mutant. Taken together, our work revealed a previously unrecognized chromatin recruitment mechanism of KAT6A, offering a new perspective on the role of KAT6A in gene regulation and human diseases.

Impaired Overall Survival in Young Patients With Acute Myeloid Leukemia and Variants in Genes Predisposing for Myeloid Malignancies.

Impaired Overall Survival in Young Patients With Acute Myeloid Leukemia and Variants in Genes Predisposing for Myeloid Malignancies.

Significance of Gene Diagnosis in Acute Myeloid Leukemia with the Emergence of New Molecular Target Drug Treatment

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic malignancy accompanied by impaired differentiation and autonomous proliferation of hematopoietic stem cells. Standard induction therapy results in first complete remission among 70% of patients with AML; however, approximately half of these patients relapse and become refractory. Allogeneic hematopoietic cell transplantation is a useful treatment for relapsed and refractory cases. However, transplantation-related mortality is approximately 20%, which is not a low value, and quality of life after transplantation decreases. Therefore, there is a need to stratify the prognosis of each patient and implement this treatment appropriately. Owing to recent advances in genome analysis technology, many gene mutations involved in onset and recurrence of AML have been discovered. These abnormalities and mutations not only have clinical application as prognostic factors and minimal residual disease markers, but they may also contribute to novel molecular targeted drug development. Many new drugs such as first-generation FMS-like tyrosine kinase 3 (FLT3), isocitrate dehydrogenase 1 and 2 (IDH1/2), and B cell lymphoma 2 (BCL2) inhibitors have been developed in the West. In addition, the second-generation FLT3 inhibitors gilteritinib and quizartinib were developed in Japan, and treatment outcomes for patients with AML have improved. However, there is still a large disparity in drug availability between the West, and Japan. As a result, treatment guidelines in the West cannot be applied in the clinical setting in Japan. In this study, we assessed the molecular target drug treatment by gene diagnosis for treatment of AML patients.

Integrated flow cytometry and sequencing to reconstruct evolutionary patterns from dysplasia to acute myeloid leukemia

AbstractClonal evolution in acute myeloid leukemia (AML) originates long before diagnosis and is a dynamic process that may affect survival. However, it remains uninvestigated during routine diagnostic workups. We hypothesized that the mutational status of bone marrow dysplastic cells and leukemic blasts, analyzed at the onset of AML using integrated multidimensional flow cytometry (MFC) immunophenotyping and fluorescence-activated cell sorting (FACS) with next-generation sequencing (NGS), could reconstruct leukemogenesis. Dysplastic cells were detected by MFC in 285 of 348 (82%) newly diagnosed patients with AML. Presence of dysplasia according to MFC and World Health Organization criteria had no prognostic value in older adults. NGS of dysplastic cells and blasts isolated at diagnosis identified 3 evolutionary patterns: stable (n = 12 of 21), branching (n = 4 of 21), and clonal evolution (n = 5 of 21). In patients achieving complete response (CR), integrated MFC and FACS with NGS showed persistent measurable residual disease (MRD) in phenotypically normal cell types, as well as the acquisition of genetic traits associated with treatment resistance. Furthermore, whole-exome sequencing of dysplastic and leukemic cells at diagnosis and of MRD uncovered different clonal involvement in dysplastic myelo-erythropoiesis, leukemic transformation, and chemoresistance. Altogether, we showed that it is possible to reconstruct leukemogenesis in ∼80% of patients with newly diagnosed AML, using techniques other than single-cell multiomics.

Abstract 5065: Predicting response to chemotherapy in a mouse model of acute myeloid leukemia

Abstract We have previously shown that state transition theory is applicable to acute myeloid leukemia (AML) to predict disease evolution. This mathematical model represents AML evolution from health to disease as a state-transition of the transcriptome represented as a particle undergoing Brownian motion in a double-well quasi-potential, where critical points define states of perturbed hematopoiesis (c1), transition to AML (c2), and overt AML (c3). We successfully demonstrated that the state transition model predicted AML onset. We now test the applicability of AML state-transition model to predict disease response to chemotherapy. To this end, we performed a time-series experiment using the conditional Cbfb-MYH11 (CM) knock-in (Cbfb+/56M/Mx1Cre) mouse model recapitulating inv(16) AML. CM leukemic mice were treated with a combination of cytarabine (50mg/kg/day; 5 days) and daunorubicin (1.5mg/kg/day, 3 days) after detection of overt leukemia which is monitored by circulating leukemia blast (cKit+ &amp;gt; 20%) to model the 7+3 standard of care treatment for newly diagnosed AML. A total of 110 peripheral blood samples from 7 CM mice were collected weekly before, during, and following chemotherapy and subjected to RNA-sequencing. The singular value decomposition was used to construct the transcriptome state-space and identified dynamics of AML (c3) after administration of chemotherapy treatment. Gene expression profiles following treatment revealed dynamics consistent with the state-transition model, with the transcriptome particle moving from leukemia (c3) towards a state of perturbed hematopoiesis (c1), before eventual relapse, re-crossing the transition point (c2) back to overt AML (c3). All 7 CM mice achieved a partial response, defined as the transcriptome particle crossing the c2 critical point, with a mean time to relapse of 5 weeks, defined as the time of the first observation after the particle crosses back over c2 towards the leukemic state c3. Mean arrival time analysis was used to accurately predict the extent of response, defined by the transcriptome particle in the state-space, and the time to relapse. We successfully applied state-transition mathematical model to predict treatment response and the time to relapse in all CM mice, confirming the applicability of the model to post-chemo therapy disease dynamics. This predictive model has implications to improve therapeutic strategies by targeting transcriptome state-transition critical points in human AML. Citation Format: Lisa Uechi, Sergio Branciamore, David E. Frankhouser, Denis O'Meally, Lianjun Zhang, Ying-Chieh Chen, Man Li, Guido Marcucci, Ya-Huei Kuo, Russell Rockne. Predicting response to chemotherapy in a mouse model of 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 5065.

3211 – MULTIOMIC ANALYSIS OF BONE MARROW HEMATOPOIETIC STEM/PROGENITOR CELLS REVEALS A POTENTIAL EARLY STAGE OF MALIGNANCY OF CLONAL HEMATOPOIESIS OF INDETERMINATE POTENTIAL

Clonal hematopoiesis of indeterminate potential (CHIP) increases the risk to develop hematological cancers such as acute myeloid leukemia (AML) tenfold. Since AML has been shown to originate from a hematopoietic stem/progenitor cell (HSPC) we compared the transcriptional landscape of bone marrow HSPCs from a healthy donor and a CHIP carrier (mutations in DNMT3A, 1.1% VAF and TET2, 5.3% VAF) by performing single cell RNA and ATAC sequencing. Cell frequency analysis of HSCP subpopulations, defined by analysis of conserved genes in clusters after dimensional reduction, showed that the proportion of HSC was twofold higher in the CHIP donor whereas the lymphocyte progenitor and pre-B cells compartment was fourfold lower, indicating an overall myeloid bias in the CHIP donor. Gene Ontology analysis of differentially expressed genes in the HSC compartment showed an upregulation of genes involved in metabolic processes in CHIP HSCs, among which genes that are associated with the onset of AML such as MEIS1 and PBX1. Furthermore, the expression levels of genes that are involved in niche retention, such as CD44 and VLA-4 were significantly higher in CHIP HSCs, suggesting alterations in the architecture of CHIP bone marrow. In addition to differences in gene expression, chromatin accessibility analysis showed unique genomic peaks in the CHIP HSC. Analysis of these sequences for binding motifs of transcription factors showed an overabundance of GATA, CEBP and RUNX1 motifs, all believed to play major roles in the onset of AML. The transcriptional landscapes of HSPC from healthy and CHIP donors show distinct differences, indicating that even the absence of signs of hematological disease, changes associated with a possible onset of malignancy can already be observed and provide possible targets for early intervention. Clonal hematopoiesis of indeterminate potential (CHIP) increases the risk to develop hematological cancers such as acute myeloid leukemia (AML) tenfold. Since AML has been shown to originate from a hematopoietic stem/progenitor cell (HSPC) we compared the transcriptional landscape of bone marrow HSPCs from a healthy donor and a CHIP carrier (mutations in DNMT3A, 1.1% VAF and TET2, 5.3% VAF) by performing single cell RNA and ATAC sequencing. Cell frequency analysis of HSCP subpopulations, defined by analysis of conserved genes in clusters after dimensional reduction, showed that the proportion of HSC was twofold higher in the CHIP donor whereas the lymphocyte progenitor and pre-B cells compartment was fourfold lower, indicating an overall myeloid bias in the CHIP donor. Gene Ontology analysis of differentially expressed genes in the HSC compartment showed an upregulation of genes involved in metabolic processes in CHIP HSCs, among which genes that are associated with the onset of AML such as MEIS1 and PBX1. Furthermore, the expression levels of genes that are involved in niche retention, such as CD44 and VLA-4 were significantly higher in CHIP HSCs, suggesting alterations in the architecture of CHIP bone marrow. In addition to differences in gene expression, chromatin accessibility analysis showed unique genomic peaks in the CHIP HSC. Analysis of these sequences for binding motifs of transcription factors showed an overabundance of GATA, CEBP and RUNX1 motifs, all believed to play major roles in the onset of AML. The transcriptional landscapes of HSPC from healthy and CHIP donors show distinct differences, indicating that even the absence of signs of hematological disease, changes associated with a possible onset of malignancy can already be observed and provide possible targets for early intervention.

Acute myeloid leukemia onset with bilateral retinal hemorrhages

Acute myeloid leukemia onset with bilateral retinal hemorrhages

C11orf21, a novel RUNX1 target gene, is down-regulated by RUNX1-ETO

The fusion protein RUNX1-ETO is an oncogenic transcription factor generated by t(8;21) chromosome translocation, which is found in FAB-M2-type acute myeloid leukemia (AML). RUNX1-ETO is known to dysregulate the normal RUNX1 transcriptional network, which should involve essential factors for the onset of AML with t(8;21). In this study, we screened for possible transcriptional targets of RUNX1 by reanalysis of public data in silico, and identified C11orf21 as a novel RUNX1 target gene because its expression was down-regulated in the presence of RUNX1-ETO. The expression level of C11orf21 was low in AML patient samples with t(8;21) and in Kasumi-1 cells, which carry RUNX1-ETO. Knockdown of RUNX1-ETO in Kasumi-1 cells restored C11orf21 expression, whereas overexpression of RUNX1 up-regulated C11orf21 expression. In addition, knockdown of RUNX1 in other human leukemia cells without RUNX-ETO, such as K562, led to a decrease in C11orf21 expression. Of note, the C11orf21 promoter sequence contains a consensus sequence for RUNX1 binding and it was activated by exogenously expressed RUNX1 based on our luciferase reporter assay. This luciferase signal was trans-dominantly suppressed by RUNX1-ETO and site-directed mutagenesis of the consensus site abrogated the reporter activity. This study demonstrated that C11orf21 is a novel transcriptional target of RUNX1 and RUNX1-ETO suppressed C11orf21 transcription in t(8;21) AML. Thus, through this in silico approach, we identified a novel transcriptional target of RUNX1, and the depletion of C11orf21, the target gene, may be associated with the onset of t(8;21) AML.