Acute Leukemia Research Articles

Diagnosis and typing of leukemia using a single peripheral blood cell through deep learning

AbstractLeukemia is highly heterogeneous, meaning that different types of leukemia require different treatments and have different prognoses. Current clinical diagnostic and typing tests are complex and time‐consuming. In particular, all of these tests rely on bone marrow aspiration, which is invasive and leads to poor patient compliance, exacerbating treatment delays. Morphological analysis of peripheral blood cells (PBC) is still primarily used to distinguish between benign and malignant hematologic disorders, but it remains a challenge to diagnose and type these diseases solely by direct observation of peripheral blood(PB) smears by human experts. In this study, we apply a segmentation‐based enhanced residual network that uses progressive multigranularity training with jigsaw patches. It is trained on a self‐built annotated dataset of 21,208 images from 237 patients, including five types of benign white blood cells(WBCs) and eight types of leukemic cells. The network is not only able to discriminate between benign and malignant cells, but also to typify leukemia using a single peripheral blood cell. The network effectively differentiated acute promyelocytic leukemia (APL) from other types of acute myeloid leukemia (non‐APL), achieving a precision rate of 89.34%, a recall rate of 97.37%, and an F1 score of 93.18% for APL. In contrast, for non‐APL cases, the model achieved a precision rate of 92.86%, but a recall rate of 74.63% and an F1 score of 82.75%. In addition, the model discriminates acute lymphoblastic leukemia(ALL) with the Ph chromosome from those without. This approach could improve patient compliance and enable faster and more accurate typing of leukemias for early diagnosis and treatment to improve survival.

FlowDiff: a simple, flow cytometry-based approach for performing a leukocyte differential count.

To overcome the challenges of a manual leukocyte differential count, we have developed FlowDiff, an 8-colour, single tube flow cytometry panel, and investigated whether it could potentially replace the manual differential in our laboratory. The instrument was set up in accordance with the EuroFlow settings, and the protocol comprised a stain-lyse no wash process, taking approximately 30 min of working time, without the addition of a toxic lysis reagent. We found a very good correlation for all leukocyte populations between FlowDiff and the Sysmex XN analyzer in 80 normal, non-flagged samples. In addition, FlowDiff showed a very good correlation with manual differential in 168 abnormal samples, as well as a high diagnostic accuracy. FlowDiff correctly identified all samples with acute leukemia (N = 13) and differentiated all B-lymphomas (N = 49) in samples with lymphocytosis. Moreover, FlowDiff detected an additional five samples with B-lymphocytosis without any prior hematological malignancy, which turned out to be a B-lymphoma. Our data suggest that FlowDiff, our 8-colour flow cytometry-based differential, is comparable to, and can successfully substitute the manual differential.

Abstract A017: IGF2BP1 is essential for ETO2/GLIS2 leukemogenesis via stabilization of primitive myeloid transcriptional programs

Abstract Certain subtypes of pediatric leukemia continue to have dismal cure rates despite advances in targeted therapy. One such disease is acute megakaryoblastic leukemia (AMKL) with a rearrangement of ETO2/GLIS2 (also known as CBFA2T3/GLIS2.) Based on our preliminary data utilizing CRISPR/Cas9 screens in vivo in patient-derived xenografts, we have nominated IGF2BP1 as a potential vulnerability in ETO2/GLIS2-rearranged AMKL. For our discovery phase, we utilized a patient-derived E2G2r AMKL which successfully engrafts in NSGS mice. We created a stable Cas9-expressing subclone using lentiviral transduction followed by FACS enrichment and limiting dilution transplants. We then generated a custom library targeting the top 100 overexpressed genes between E2G2r leukemias versus other leukemia PDXs which we previously characterized, including eight transcription factors with motifs enriched in differentially expressed genes, as well as eight pan-essential positive controls. Mice were sacrificed at humane endpoints, and spleens and bone marrow underwent PCR amplification for gRNA sequences followed by sequencing and analysis with MAGeCK-VISPR (Wei L, et al. Genome Biol 2015). We observed the dropout of 38 genes with a false discovery rate < 0.25, including all positive control gRNAs. Among the most significantly depleted genes was the RNA binding protein IGF2BP1. We subsequently characterized phenotypic changes using shRNAs in the E2G2r cell line M07e and ETO2/GLIS2-transformed CD34+ umbilical cord blood cells. Notably, we observed impaired cell growth with IGF2BP1 knockdown in both cell lines as well as the parent xenograft. We also generated M07e lines expressing a degron (FKBPF36V, aka “dTAG”, Nabet et al. Nat Chem Biol 2018) knocked in at the endogenous IGF2BP1 locus and observed similar phenomena. Through flow cytometry of M07e following IGF2BP1 depletion, we observed changes in the leukemic immunophenotype as well, suggesting perturbation of megakaryocyte differentiation potential. To determine the mechanism underlying our observations, we performed enhanced UV crosslinking with IP and sequencing (Blue SM et al. Nat Protoc 2022) against both IGF2BP1 and m6A residues based on the putative role of IGF2BP1 as an m6A reader. We observed binding of IGF2BP1 predominantly in 3’UTRs, as well as the presence of m6A-modified bases at IGF2BP1 binding sites. Gene ontology (GO) analysis of IGF2BP1-bound transcripts demonstrated enrichment for GO terms pertaining to embryonic hematopoiesis and megakaryocyte maturation/development. We further evaluated global changes in gene expression following IGF2BP1 loss, and performed RNAseq 72 hours following shRNA-mediated knockdown. GO analysis showed that downregulated genes were highly enriched for E2F-mediated transcriptional programs. We hypothesize that IGF2BP1 regulates myeloid developmental transcripts essential for E2G2r leukemogenesis, likely via stabilization in an m6A-dependent fashion, and thus may represent a selective therapeutic target. Work is ongoing to further elucidate this proposed mechanism. Citation Format: Jason Clark, Mark Wunderlich, Jing Chen, Daniel Starczynowski, Lynn Lee. IGF2BP1 is essential for ETO2/GLIS2 leukemogenesis via stabilization of primitive myeloid transcriptional programs [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr A017.

Improved identification of clinically relevant Acute Leukemia subtypes using standardized EuroFlow panels versus non-standardized approach.

Rare acute leukemia (AL) components or subtypes such as blastic plasmacytoid dendritic cell neoplasm (BPDCN) or early T-cell precursor acute Lymphoblastic Leukemia (ETP-ALL) can be difficult to detect by routine flow cytometry due to their immunophenotypes overlapping with other poorly differentiated AL. We hypothesized that using standardized EuroFlow™ Consortium approach could better diagnose such entities among cases that previously classified as acute myeloid leukemia (AML)-M0, AML with minimal differentiation, AML with myelodysplasia-related changes without further lineage differentiation, and AL of ambiguous lineage. In order to confirm this hypothesis and assess whether these AL subtypes such as BPDCN and ETP-ALL had previously gone undetected, we reanalyzed 49 banked cryopreserved sample cases using standardized EuroFlow™ Consortium panels. We also performed target sequencing to capture the mutational commonalities between these AL subtypes. Reanalysis led to revised or refined diagnoses for 23 cases (47%). Of these, five diagnoses were modified, uncovering 3 ETP-ALL and 2 typical BPDCN cases. In 12 AML cases, a variable proportion of immature plasmacytoid dendritic cell and/or monocytic component was newly identified. In one AML case, we have identified a megakaryoblastic differentiation. Finally, in five acute lymphoblastic leukemia (ALL) cases, we were able to more precisely determine the maturation stage. The application of standardized EuroFlow flow cytometry immunophenotyping improves the diagnostic accuracy of ALs and could impact treatment decisions.

Disease Pattern and Clinical Profile of Admitted Patients in Pediatric Intensive Care Unit of a Tertiary Care Hospital in Chattogram

Background: The care of critically ill children remains one of the most demanding and challenging aspects in the field of pediatrics. Pediatric Intensive Care Unit (PICU) aims at promoting early intervention and quality care for achieving good results and better prognosis. With the advancement in intensive care facilities, there is a dramatic increase in survival of critically ill children. It is usually offered to patients whose condition is potentially reversible and who have a good chance of surviving with intensive care support. Since these patients are critically ill, the outcome of intervention is sometimes difficult to predict. This study is aimed to assess the clinical profile of critically ill children presenting to the pediatric emergency room and then their admission pattern to the pediatric intensive care unit. This may help to assist intensivist and planners to pay due attention for better utilization of healthcare facilities. Better understanding leads to better management. Materials and methods: This is a cross-sectional study, conducted at the Pediatric Intensive Care Unit of Chattagram Maa Shishu-O-General Hospital from January to June of 2019. A total of 100 admitted cases were recruited and their symptoms and signs, clinical findings and diagnosis were recorded accordingly. All the patients were aged within 28 days to below 14 years. Results: Among the 100 study subjects, 66 were male and 34 were female. The mean age was 1.73 ± 2.6 years. Most children were admitted with respiratory distress though there were other accompanying signs and symptoms. The symptoms subjects commonly presented with were respiratory distress(76%), tachycardia(70%), lung crepitations (69%), tachypnea (68%), fever (67%), lethargy (59%), feeding difficulty (59%), convulsions (48%), low blood pressure (24%), hepatomegaly (19%), cyanosis (14%), edema (6%), congenital anomalies (4%). The clinical findings that led them to PICU admissions were bronchiolitis (4%), bronchopneumonia (42%), respiratory failure of any cause (4%), heart failure (7%), cyanotic congenital heart disease (8%), acyanoticcongenital heart disease (7%), meningitis (13%), encephalitis (12%), cerebral palsy with seizure disorder (21%), status epilepticus (2%), Gullein Barre Syndrome (3%), hyponatremic dehydration (6%), acute kidney injury (2%), acute leukemia (2%).In this study, neurological disorders such as cerebral palsy with seizure disorder and encephalitis and meningitis were among the top five in the diagnosis list. Septicemia with shock was also a very common finding promoting admission to PICU. GCS was found to be significantly lower in the above-mentioned disease conditions making admission to the ICU quite plausible. Conclusion: Children admitted to PICU may have multiple clinical findings and disease patterns. However, some diseases predominate the patient population admitted to PICU.The knowledge of clinical spectrum and epidemiological profile of critically ill children plays a significant role in the planning of health policies that would mitigate various factors related to the evolution of diseases prevalent in these sectors. Descriptive epidemiology focuses on identifying and reporting the pattern and frequency of events related to the health of a population. IAHS Medical Journal Vol 7(1), June 2024; 44-49

Acute promyelocytic leukemia as a differential diagnosis of Crimean-Congo hemorrhagic fever

Acute promyelocytic leukemia as a differential diagnosis of Crimean-Congo hemorrhagic fever

Radiation Therapy for Cutaneous Blastic Plasmacytoid Dendritic Cell Neoplasm: A Case Report and Review of the Literature

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a clinically aggressive hematologic malignancy derived from plasmacytoid dendritic cells. It commonly presents as cutaneous lesions. To date, no standard treatment protocol for BPDCN exists. Traditionally treated similarly to acute leukemia or lymphoma, its prognosis remains poor. Radiation therapy is employed for isolated skin lesions, for patients that are ineligible for chemotherapy due to age or comorbidities and for post-chemotherapy recurrence. However, very limited reports are available on radiotherapy for BPDCN. We present a case involving a 94-year-old BPDCN patient treated with radiation therapy, highlighting an atypical situation of two separate radiotherapy sessions with different dosages for isolated skin lesions. Initially, 45 Gy was administered in 15 fractions (45 Gy/15 Fr), followed by a second session of 30 Gy in 10 fractions (30 Gy/10 Fr) after disease recurrence. This case is unique in detailing radiation therapy for the exceedingly rare BPDCN, particularly dose fractionation. The findings indicate that 45 Gy/15 Fr can provide adequate local control, while even a lower dose of 30 Gy/10 Fr may be effective. This case report contributes to the limited literature by proposing potential therapeutic approaches and dosage guidelines to refine future BPDCN treatment protocols.

Current insights on m6A RNA modification in acute leukemia: therapeutic targets and future prospects

RNA modification is the critical mechanism for regulating post-transcriptional processes. There are more than 150 RNA modifications reported so far, among which N6-Methyladenosine is the most prevalent one. M6A RNA modification complex consists of ‘writers’, ‘readers’ and ‘erasers’ which together in a group catalyze, recognize and regulate the methylation process of RNA and thereby regulate the stability and translation of mRNA. The discovery of erasers also known as demethylases, revolutionized the research on RNA modifications as it revealed that this modification is reversible. Since then, various studies have focused on discovering the role of m6A modification in various diseases especially cancers. Aberrant expression of these ‘readers’, ‘writers’, and ‘erasers’ is found to be altered in various cancers resulting in disturbance of cellular homeostasis. Acute leukemias are the most common cancer found in pediatric patients and account for 20% of adult cases. Dysregulation of the RNA modifying complex have been reported in development and progression of hematopoietic malignancies. Further, targeting m6A modification is the new approach for cancer immunotherapy and is being explored extensively. This review provides detailed information about current information on the role of m6A RNA modification in acute leukemia and their therapeutic potential.

Global research trends in traditional Chinese medicine therapy for acute leukemia: a comprehensive visualization and bibliometric analysis

ABSTRACT Background: The application of traditional Chinese medicine (TCM) therapy to acute leukemia has been intensively investigated. However, the bibliometric analysis in this field has not been performed. This bibliometric study aimed to comprehensively analyze the research trends and active areas of TCM therapy for acute leukemia from 2000 to 2023. Methods: We searched articles and reviews published between 2000 and 2023 that discussed TCM in acute leukemia from the Web of Science Core Collection (WoSCC). Knowledge mapping and bibliometric analysis were conducted using VOSviewer, CiteSpace software, and R-bibliometrix. Results: A total of 1,099 articles were included, with China, the United States, and Korea contributing the most papers. Most papers were published in the Journal of Ethnopharmacology. Meanwhile, China saw a steady increase in the number of publications. The three leading institutions that made outstanding contributions were the China Medical University, the Chinese Academy of Sciences, and the China Academy of Chinese Medical Sciences. Efferth Thomas, Liu Wei, and Liu Jie were the top three productive authors, with Efferth T receiving the most co-citations. The most frequently cited reference was Shen ZX (1997). In the analysis of keywords co-occurrence, ‘survival,’ ‘risk factors,’ ‘nanoparticles,’ and ‘metabolism’ are the active research topics. Conclusion: This bibliometric study provides researchers with a comprehensive overview and significant value in understanding the development of TCM in acute leukemia treatment.

Mixed-phenotype leukemia with TCF3::ZNF384 fusion presenting as an isolated mediastinal mass.

Acute leukemia with TCF3::ZNF384 is a distinct type of acute leukemia that present most commonly as B-acute lymphoblastic leukemia or mixed-phenotype acute leukemia (B/myeloid). We report the first case of TCF3::ZNF384 mixed-phenotype leukemia presenting as isolated extramedullary disease in the mediastinum. Diagnosis using RNA-sequencing and whole genome sequencing on the primary issue is illustrated.

DDDR-36. LOOKING AROUND THE CORNER: PREDICTING SYNERGY OR FUTILITY TO COMBINATION OF ARSENIC TRIOXIDE PLUS MNK1 INHIBITORS IN GLIOBLASTOMA

Abstract Glioblastoma (GBM) is the most prevalent type of malignant tumor within the central nervous system. The five-year survival rate of 6.8% highlights the challenges inherent to exceptionally heterogeneous tumors that grow behind the blood brain barrier. The protracted dearth in new agents approved for GBM begs innovative strategies by which to approach clinical trials. Arsenic trioxide (ATO) is the standard of care for myelodysplasia and relapsed or refractory acute promyelocytic leukemia. ATO has an idiopathic mechanism of action, but has shown beneficial effects in other solid tumors. Across six GBM PDX models a 10-fold difference in sensitivity manifests, indicative of underlying innate sensitivity or resistance to ATO. This resistance has been previously shown to correlate with MNK1 expression. In order to sensitize PDX models innately resistant to ATO we utilized combination therapy with the MNK1 inhibitor AUM001 (AUM Biosciences, Singapore; NCT05462236). In high MNK1 expressing PDX models the combination of ATO + AUM001 induces cytotoxic synergy. Additionally, when narrowly measuring effects on the glioma stem cell population (ELDA), strong synergy was observed in models, including those failing to show such using bulk cell population viability assay. GSEA analysis revealed differential high enrichment in gene sets related to oxidative phosphorylation in the models failing to show synergy. Such criteria could act as exclusionary criteria (futility) for clinical trials by excluding such putative non-responder patients from the treatment population. Synergistic models showed enrichment in gene sets related to epithelial/mesenchymal transition, skeletal and sensory organ development, and regionalization. These signatures could serve to prioritize patient inclusion in clinical trials of ATO in combination with AUM001, enriching the fraction of responsive glioma patients. Such molecular signatures may bring precision medicine in early-stage clinical trials to advance promising new agents and combinations of agents to full FDA approval.

Clinical Significance of Complement and Coagulation Cascades Genes for Patients With Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) is the second most common acute leukemia in adults and the 5-year survival remains low. We analyzed the gene expression profiles of the complement and coagulation cascades pathway (CCCP) in 998 bone marrow (BM) and 122 peripheral blood (PB) samples of ALL patients and healthy individuals obtained from the TCGA database and evaluated their clinical significance in terms of being diagnostic and prognostic biomarkers. We identified 18 CCCP genes (SERPINA1, C5AR1, F5, CD55, PLAUR, C3AR1, THBD, CD59, PLAU, VWF, CFD, F13A1, C1QA, C1QB, C1QC, A2M, SERPINE1 and CR2) differentially expressed in the BM samples of ALL patients compared to healthy individuals. The expression levels of CD55, F13A1 and CR2 in BM were linked with the overall survival of ALL patients. While in PB only 11 CCCP genes (e.g., SERPINA1, C5AR1, F5, PLAUR, C3AR1, THBD, CFD, F13A1, C1QA, SERPINE1, and CR2) were differentially expressed and F13A1 was significantly associated with ALL patient survival. Machine learning enabled us to predict ALL using the CCCP genes and the accuracy can reach 0.9701 and 0.9167 using the BM and PB, respectively. Furthermore, using single-cell RNA sequencing, we found that the differential expression of CCCP genes was found with diversity in the BM-derived immune cells of ALL patients. Our findings suggest that the CCCP genes may play a key role in the progression of ALL and can be used as potential therapeutic targets and diagnostic markers.

Anticancer effects of pomegranate-derived peptide PG2 on CDK2 and miRNA-339-5p-mediated apoptosis via extracellular vesicles in acute leukemia

Acute leukemia has rapid onset and severe complications. Anticancer peptides from natural sources have demonstrated efficacy in eliminating various cancers through apoptosis signaling pathways. Additionally, extracellular vesicles containing microRNAs play pivotal roles in promoting tumorigenesis. Therefore, this study aimed to investigate the impact of PG2, a pomegranate peptide that regulates extracellular vesicles, on the induction of acute leukemia cell apoptosis. NB4 and MOLT-4 leukemia cell lines were treated with PG2 alone or in combination with daunorubicin to assess cell viability using the MTT assay. Extracellular vesicles were extracted from PG2-treated NB4 and MOLT-4 cells. Bioinformatic tools were utilized to predict target proteins and microRNAs, following which mRNA and protein expression were determined by using RT‒qPCR and western blotting, respectively. PG2 significantly reduced the viability of NB4 and MOLT-4 cells. Furthermore, the combination of PG2 with daunorubicin had a synergistic effect on NB4 and MOLT-4 cells. Subsequent treatment with PG2 or PG2-treated extracellular vesicles decreased CDK2 expression while increasing microRNA-339-5p and caspase-3 expression in NB4 and MOLT-4 cells. Our findings revealed that the anticancer activity of PG2 through the CDK2/miR-339-5p/caspase-3 pathway is mediated by extracellular vesicles, ultimately inducing apoptosis. PG2 holds promise as a potential antileukemic drug.

Pioneering sustainable treatment delivery in childhood leukemia through synchronous telemedicine-A pilot study.

Cancer care places a heavy economic burden on families and health systems, driven by high treatment costs, lengthy hospital stays, and the necessity for extensive travel to specialized facilities. To address this challenge, an integrated health care network (IHCN) was implemented for maintenance treatment in acute leukemia. The IHCN encompassed outpatient services provided by local physicians and synchronous telemedicine consultation with pediatric oncologists. This study included twenty-two pediatric patients (eleven [50.0%] females; twenty [90.9%] with B-ALL and two [9.1%] with AML). The IHCN was offered to all rural patients (n = 17) with a one-way driving distance more than 30 km, while urban patients (n = 5) received regular cancer care. Throughout the study, rural patients had a total of 510 routine clinical visits, with 367 (72%) conducted through the IHCN. Physical examinations revealed similar frequency of new abnormal findings for urban and rural patients (22.4% vs. 17.8%; p = .31). Laboratory tests indicated no significant difference in the frequency of abnormal values for various parameters between both groups. Similarly, there was no discrepancy of drug modifications or interruption in maintenance therapy between the two settings (p = .85). Moreover, patients' health-related quality of life remained within the normative range, and user satisfaction with the IHCN was notably high. The implementation of the IHCN resulted in savings of 70,158 km, 950 h of travel, and 12,277 kg CO2 emissions. This pilot study underscores the efficacy of a telemedicine-based IHCN, ensuring safety, quality of care, cost reduction, and satisfaction for both families and health care providers in pediatric leukemia management.

The relationship between seasonality and the diagnosis of acute leukaemia in central South Africa

Background: Leukaemias are haematological malignancies resulting from the abnormal clonal proliferation of haematopoietic precursors. Their incidence is influenced by various environmental and genetic factors.Aim: With this retrospective descriptive study, the authors aimed to explore the possible influence of seasonality on the type and number of acute leukaemia (AL) diagnoses made at the Universitas Academic Hospital (UAH) National Health Laboratory Service (NHLS) from 01 January 2018 to 31 December 2021.Setting: Universitas Academic Hospital, Bloemfontein, South Africa.Methods: Archived laboratory reports of all patients diagnosed with lymphoid and myeloid during the study period were included. Patients’ age, sex, ethnicity, final diagnosis and date of diagnosis were recorded. Information was pseudonymised to maintain confidentiality. A descriptive statistical analysis was performed to explore the possible influence of seasonality on the number of cases and type of leukaemia diagnosed.Results: In all, 249 patients were included. Acute myeloid leukaemia (AML) was the most common AL subtype (n = 117; 47.0%). Over the 4-year study period, all AL subtypes were more frequently diagnosed in summer (n = 131; 52.6%). However, the monthly number of AL diagnoses was relatively consistent over the 4 years for all subtypes (p = 0.7603), with consistent peaks of AML cases during January and February (summer) and May (autumn).Conclusion: No statistically significant association between the different seasons and AL diagnosis was noted.Contribution: Further studies using a larger study population and a wider geographical area, conducted over a more extended period, might affect the observations made in this study

Multiplexed gastrointestinal PCR panels for the evaluation of diarrhea in patients with acute leukemia.

To better delineate multiplexed gastrointestinal polymerase chain reaction (PCR) panel (MGPP) diagnostic and therapeutic stewardship for patients undergoing treatment for acute leukemia including indications and benefits of testing, optimal timing, and interpretation of results. We retrieved all MGPP ordered on 662 consecutive patients admitted with newly diagnosed acute leukemia between June 2015 and May 2024. Regional referral center for acute leukemia. Fifty-one (17%) of 305 MGPP obtained on the 198 patients who underwent testing identified at least one and 4 (1%) more than one diarrheagenic pathogen. The probability of a positive result was greater if obtained as an outpatient [20/52(38%)], but was not related to type of leukemia, sex, or age. Among the positive results, the pathogens identified included Clostridioides difficile (78% of tests), norovirus (16%), diarrheagenic Escherichia coli (6%), adenovirus 40/41 (4%), and Giardia lamblia (4%). The results of 30 of the 305 tests resulted in a change in treatment (28 C. difficile, 2 G. lamblia). For the MGPP C. difficile results with an accompanying toxin determination, this included treatment following 16/19 tests with a positive toxin result and 11/19 with a negative. Actionable results other than C. difficile were rarely seen in the inpatient population. MGPP testing is most useful when administered as an outpatient and of little benefit for inpatients with hospital-onset diarrhea. Since MGPP is sensitive and does not distinguish between colonization and causes of diarrhea, caution is needed in interpretation of results, especially for toxin-negative C. difficile.

Enhancing Gene Delivery in NB-4 Cells: Overcoming Transduction and Selection Challenges

Efficient gene transduction and cell viability are critical factors in genetic manipulation for research and therapeutic purposes. In this study, we explored the challenges associated with transducing the NB-4 cell line, a well-established model for acute promyelocytic leukemia (APL), using lentiviral vectors. While the initial transduction efficiency in NB-4 cells reached approximately 30%, we observed a significant decrease in cell viability, a phenomenon not observed in other acute leukemia cell lines such as THP-1 cells. We identified that this toxicity could be mitigated by purifying viral particles through ultracentrifugation or polyethylene glycol (PEG) precipitation, indicating that toxic substances, potentially secondary metabolites released by HEK293, could be responsible for the cell death. Nevertheless, cell selection by puromycin was still ineffective; crucially, we discovered that the human phosphoglycerate kinase (hPGK) promoter, commonly used in the PLKO1 vector, may become silenced in NB-4 cells, preventing effective selection with puromycin. By replacing the hPGK promoter with the elongation factor-1 alpha (EF1α) promoter, we successfully achieved high transduction efficiency and robust selection, demonstrating the potential for this modified vector system to facilitate genetic studies in APL models. These findings provide important insights into optimizing gene transduction protocols not only for NB-4 cells but also for other challenging cell lines, offering a refined approach for gene delivery and selection in cell models.

Arsenic trioxide neurotoxicity in acute promyelocytic leukemia patients: a single center experience

Arsenic trioxide neurotoxicity in acute promyelocytic leukemia patients: a single center experience

Isolated peripheral neuroleukemiosis mimicking Guillain‐Barré syndrome in an adolescent with relapsed B‐lymphoblastic leukemia

AbstractBackgroundNeuroleukemiosis, leukemic infiltration of the peripheral nervous system (PNS), is rare. Very few cases of isolated neuroleukemiosis, PNS leukemic infiltration without leukemia blasts in the blood or bone marrow, have been reported in pediatrics. Most cases have occurred in adults with acute myelogenous leukemia (AML) in remission who presented with peripheral neuropathy. We describe a pediatric patient presenting with isolated neuroleukemiosis mimicking Guillain‐Barré syndrome.Patient descriptionA 15‐year‐old boy presented 1 month after IVIG treatment for Guillain‐Barre syndrome with worsening ataxia and weakness. He had a past medical history of B‐cell acute lymphoblastic leukemia (B‐ALL) in remission for 8 years. Examination revealed distal greater than proximal weakness of the bilateral lower extremities, areflexia at the Achilles tendon, and 1+ patellar and upper extremity reflexes. Initial magnetic resonance imaging (MRI) of the brain and spine were normal, and lumbar puncture revealed increased protein with uninterpretable white count due to excessive red blood cells. Repeat MRI brain and spinal cord showed extensive enlargement of all nerve roots and enhancement of the cauda equina and portions of cranial nerve VII bilaterally. Repeat lumbar puncture with cytology revealed leukemic blast cells. Blasts were not detected in peripheral blood smear or by flow cytometry. Bone marrow biopsy was also free of blast cells, confirming the diagnosis of relapsed B‐ALL restricted to the nervous system.ConclusionsNeuroleukemiosis is a rare entity with typical clinical features of mono‐ or polyneuropathy. It most often occurs in adults in remission from AML. However, neuroleukemiosis should also be on the differential for pediatric patients in remission from ALL presenting with neuropathy.

Discovery of KW0113 as a First and Effective PROTAC Degrader of DNMT1 Protein.

DNA methyltransferase 1 (DNMT1) is an attractive therapeutic target for acute myelocytic leukemia (AML) and other malignancies. It has been reported that the genetic depletion of DNMT1 inhibited AML cell proliferation through reversing DNA methylation abnormalities. However, no DNMT1-targeted PROTAC degraders have been reported yet. Herein, a series of proteolysis-targeting chimera (PROTAC) degrader of DNMT1 based on dicyanopyridine scaffold and VHL E3 ubiquitin ligase ligand was developed. Among them, KW0113 (DC50=643/899 nM in MV4-11/MOLM-13 cells) exhibited optimal DNMT1 degradation. KW0113 induced DNMT1-selective degradation in a dose- and time-dependent manner through VHL engagement. Moreover, KW0113 inhibited AML cell growth by reversing promoter DNA hypermethylation and tumor-suppressor genes silencing. In conclusion, these findings proved the capability of PROTAC strategy for inducing DNMT1 degradation, demonstrated the therapeutic potential of DNMT1-targeted PROTACs. This work also provided a convenient chemical knockdown tool for DNMT1-related studies.