scholarly journals Distribution and levels of cell surface expression of CD33 and CD123 in acute myeloid leukemia

2014 ◽  
Vol 4 (6) ◽  
pp. e218-e218 ◽  
Author(s):  
A Ehninger ◽  
◽  
M Kramer ◽  
C Röllig ◽  
C Thiede ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Surface ◽  
Myeloid Leukemia ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Acute Myeloid

scholarly journals Therapeutic Targeting of Mesothelin with Chimeric Antigen Receptor Natural Killer Cell Therapy in Acute Myeloid Leukemia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1712-1712
Author(s):  
Thao T. Tang ◽  
Lindsey F Call ◽  
Sommer Castro ◽  
Cynthia Nourigat-Mckay ◽  
LaKeisha Perkins ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Surface ◽  
Nk Cells ◽  
Myeloid Leukemia ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Car T Cells ◽  
Car T ◽  
Acute Myeloid

Abstract Effective immunotherapy for acute myeloid leukemia (AML) has been limited by the lack of AML-restricted targets (expression in AML but silent in normal hematopoiesis). Current immunotherapies targeting lineage markers CD33 and CD123 (if effective) would lead to prolonged myelosuppression or myeloablation, requiring stem cell transplantation to restore hematopoiesis after treatment. In search for AML-restricted targets, we interrogated the AML transcriptome of nearly 3000 cases in pediatric and young adults and contrasting it to normal bone marrow and peripheral blood CD34+ samples. This extensive discovery effort has identified over 200 AML-restricted targets with mesothelin (MSLN) emerged as one of the highest expressing AML-restricted targets and highly enriched in the KMT2A-rearranged AML subtype. We previously validated cell surface expression of MSLN on both AML blasts and leukemic stem cells (Le et. al. 2021). We further showed efficacy targeting MSLN in AML using antibody-drug conjugates (Kaeding et. al. 2021) and chimeric antigen receptor (CAR) T cells (Le et. al. 2021). Given that natural killer (NK) cells are potent immune effector cells and generally have a more favorable toxicity profile than CAR T cells (i.e without cytokine release syndrome), we developed CAR NK cells targeting MSLN and evaluated their efficacy in AML preclinical models. From primary patient samples, we verified MSLN cell surface expression and showed high correlation between cell surface expression (mean fluorescence intensity, MFI) and transcript expression (TPM, R = 0.72, p = 2.1x10 -8, Figure 1A) . Importantly, MSLN expression was restricted to AML blasts and entirely absent in normal lymphocytes and myeloid cells in individual patients (Figure 1B, C). Having confirmed cell surface and AML-restricted expression of MSLN, we developed CAR NK cells targeting MSLN. The VH and VL sequences from immunotoxin SS1P were used to create the single-chain variable fragment domain of the standard CAR (41-BB and CD3Zeta, Figure 1D). CAR NK cells were generated by transducing NK-92 cells with the MSLN CAR construct. Cytotoxicity of CAR NK cells was evaluated against Nomo-1 AML cell line, which expresses endogenous level of MSLN; MV4;11 and Kasumi-1 cell lines engineered to express MSLN with a lentivirus construct (MV4;11 MSLN+ and Kasumi-1 MSLN+). We initially tested the target specificity of MSLN-directed CAR NK cells against MSLN-positive (Nomo-1, MV4;11 MSLN+ and Kasumi-1 MSLN+) and MSLN-negative (Nomo-1 MSLN KO, MV4;11 and Kasumi-1) cells. CAR NK cells exhibited enhanced cytolytic activity against MSLN-positive but not MSLN-negative AML cells after 12 hours of co-incubation at the indicated effector: target (E:T) ratios (Figure 1E). We next assessed the in vivo efficacy of CAR NK cells. Nomo-1 cells transduced with GFP/Luciferase were transplanted into NSG mice at 1x10 6 cells/mouse. Unmodified or CAR NK cells were infused 1 week following leukemic cell injection at 1x10 7 cells/mouse. Monitoring leukemia burden by bioluminescence IVIS imaging showed that after 4 days post NK injection, the leukemia was significantly reduced in Nomo-1 xenograft mice treated with CAR NK cells compared to mice that received unmodified NK cells (Figure 1F), suggesting highly potent anti-leukemia activity of CAR NK cells. In this study, we demonstrate that the cell surface expression of MSLN is restricted to AML blasts but is entirely silent in normal hematopoietic subsets in individual patients. Previous and current clinical trials utilizing NK-92 cells have demonstrated safety and efficacy in variety of cancers, including AML. Here, we demonstrate that NK-92 cells genetically modified with a CAR to redirect their specificity against MSLN-positive AML cells exhibit potent, target-dependent anti-leukemia activity in vitro and in vivo. These results provide compelling data to evaluate MSLN-directed CAR NK cell therapy in clinical trials for refractory/relapsed AML, especially for high-risk KMT2A-rearranged leukemia where majority of patients express MSLN at diagnosis and relapse. Figure 1 Figure 1. Disclosures Pardo: Hematologics, Inc.: Current Employment.

scholarly journals CD33_PGx6_Score Predicts Gemtuzumab Ozogamicin Response in Childhood Acute Myeloid Leukemia: A Report From the Children’s Oncology Group

2019 ◽  
pp. 1-15 ◽  
Author(s):  
Lata Chauhan ◽  
Miyoung Shin ◽  
Yi-Cheng Wang ◽  
Michael Loken ◽  
Jessica Pollard ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Surface ◽  
Clinical Response ◽  
Myeloid Leukemia ◽  
Prognostic Significance ◽  
Disease Free Survival ◽  
Gemtuzumab Ozogamicin ◽  
Cell Surface Expression ◽  
Oncology Group ◽  
Acute Myeloid

PURPOSE The US Food and Drug Administration recently announced reapproval of gemtuzumab ozogamicin (GO) for treatment of CD33-positive acute myeloid leukemia (AML), thus opening up opportunities to develop strategies for effective use of GO. In light of our recent report showing prognostic significance of CD33 splicing single nucleotide polymorphisms (SNPs), the objective of this study was to comprehensively evaluate CD33 SNPs for accurate prediction of patients with AML who are more or less likely to respond to GO. PATIENTS AND METHODS We investigated the five new CD33 SNPs (rs2455069, rs35112940, rs61736475, rs1803254, and rs201074739) for association with CD33 leukemic cell surface expression and clinical response in pediatric patients with AML enrolled in the Children’s Oncology Group AAML0531 trial. We further developed a composite CD33 pharmacogenetics (PGx) score using six CD33 SNPs (CD33_PGx6_score) for association with clinical outcome. RESULTS Four CD33 SNPs were associated with cell surface CD33 levels and clinical response in the GO versus no-GO arms. Therefore, the CD33_PGx6_score was built using directional genotype scores for the previously reported splicing SNP and five new SNPs. Patients with a CD33_PGx6_score of 0 or higher had higher CD33 expression levels compared with patients with a score of less than 0 ( P < .001). In addition, patients with a score of 0 or higher demonstrated an improved disease-free survival in the GO versus no-GO arms (62.5% ± 7.8% v 46.8% ± 8.3%, respectively; P = .008) and a reduced risk of relapse (28.3% ± 7.2% v 49.9% ± 8.4%, respectively; P < .001). No improvement from GO was observed in patients with a CD33-PGx6_score of less than 0. Consistent results were observed across the risk groups. CONCLUSION In this study, we report a composite CD33_PGx6_score using directional genotype scores of CD33 SNPs. Once validated, our findings hold promise for use of the CD33_PGx6_score to guide efficient use of GO in patients with AML. In addition, because the CD33_PGx6_score considers SNPs with varying abundance in different ethnic groups, it has potential for global application.

scholarly journals Aspartate β-Hydroxylase (ASPH) Expression in Acute Myeloid Leukemia: A Potential Novel Therapeutic Target

2021 ◽  
Vol 11 ◽  
Author(s):  
Noa G. Holtzman ◽  
Michael S. Lebowitz ◽  
Rima Koka ◽  
Maria R. Baer ◽  
Kanam Malhotra ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Transmembrane Protein ◽  
Expression Patterns ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Fluorescent Intensity ◽  
Clinical Prognosis ◽  
Disease Characteristics ◽  
Acute Myeloid

BackgroundAspartate β-hydroxylase (ASPH) is an embryonic transmembrane protein aberrantly upregulated in cancer cells, associated with malignant transformation and, in some reports, with poor clinical prognosis.ObjectiveTo report the expression patterns of ASPH in acute myeloid leukemia (AML).MethodsCell surface expression of ASPH was measured via 8-color multiparameter flow cytometry in 41 AML patient samples (31 bone marrow, 10 blood) using fluorescein isothiocyanate (FITC)-conjugated anti-ASPH antibody, SNS-622. A mean fluorescent intensity (MFI) of 10 was used as a cutoff for ASPH surface expression positivity. Data regarding patient and disease characteristics were collected.ResultsASPH surface expression was found on AML blasts in 16 samples (39%). Higher ASPH expression was seen in myeloblasts of African American patients (p=0.02), but no correlation was found between ASPH expression and other patient or disease characteristics. No association was found between ASPH status and CR rate (p=0.53), EFS (p=0.87), or OS (p=0.17).ConclusionsASPH is expressed on blasts in approximately 40% of AML cases, and may serve as a new therapeutically targetable leukemia-associated antigen.

Novel CD33 antibodies unravel localization, biology and therapeutic implications of CD33 isoforms

2020 ◽  
Author(s):  
Mohammed O Gbadamosi ◽  
Vivek M Shastri ◽  
Tiffany Hylkema ◽  
Ioannis Papageorgiou ◽  
Laura Pardo ◽  
...  
Keyword(s):  
Cell Surface ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Nucleotide Polymorphism ◽  
Minimal Cell ◽  
Single Nucleotide ◽  
Therapeutic Implications ◽  
Acute Myeloid

The aim of this study was to establish the therapeutic relevance of the CD33D2 isoform by developing novel antibodies targeting the IgC domain of CD33. Two novel IgC-targeting antibodies, HL2541 and 5C11-2, were developed, and CD33 isoforms were assessed using multiple assays in cells overexpressing either CD33FL or CD33D2 isoforms, unmodified acute myeloid leukemia (AML) cell lines and primary AML specimens representing different genotypes for the CD33 splicing single nucleotide polymorphism. CD33D2 was recognized on cells overexpressing CD33D2 and unmodified AML cell lines; however, minimal/no cell surface detection of CD33D2 was observed in primary AML specimens. Both isoforms were detected intracellularly using novel antibodies. Minimal cell surface expression of CD33D2 on primary AML/progenitor cells warrants further studies on anti-CD33D2 immunotherapeutics.

scholarly journals Enhancement of Eligibility Guidelines for Gemtuzumab Ozogamicin Therapy for Childhood Acute Myeloid Leukemia: A Report from Children's Oncology Group Protocol AAML0531

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1490-1490
Author(s):  
Laura Pardo ◽  
Lisa Eidenschink Brodersen ◽  
Jatinder K Lamba ◽  
Todd A Alonzo ◽  
Yi-Cheng Wang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Acute Myeloid Leukemia ◽  
Cell Surface ◽  
Myeloid Leukemia ◽  
Gemtuzumab Ozogamicin ◽  
Response To Therapy ◽  
Cell Surface Expression ◽  
Oncology Group ◽  
Expression Levels ◽  
Acute Myeloid

Abstract Background: CD33 is variably expressed on leukemia blasts in most patients with acute myeloid leukemia (AML). Efforts to target CD33 therapeutically have focused on gemtuzumab ozogamicin (GO), an antibody-drug conjugate delivering a DNA-damaging calicheamicin derivative. Qualification for GO therapy has been determined by expression of CD33 by multidimensional flow cytometry (MDF); patients with positive CD33 expression receive GO. Previous studies from the AAML0531 protocol demonstrated that the cell surface intensity expression of CD33, determined by MDF, predicts the response to treatment with GO, and that in part this expression is regulated by a pair of CD33 single nucleotide polymorphisms (SNPs) in linkage disequilibrium that also, independently, predict response to therapy. Patients with CC genotype for rs12459419 have lower relapse risk and improved disease-free survival, compared to CT and TT genotypes. Because GO therapy is associated with treatment-related toxicity, it is important to identify biologic variables associated with benefits and risks. To date, there has been no report associating SNP status among CD33 positive versus CD33 negative patients that could assess how the combination of these biomarkers for determining administration of GO therapy could improve patient outcomes. Objective: In an effort to determine which children would benefit most from GO treatment in future prospective pediatric AML protocols, we aimed to elucidate if CD33 SNP genotype status should be combined with quantitative CD33 cell surface antigen expression on the diagnostic leukemia cells (CD33+ versus CD33-) to determine GO eligibility, with a retrospective analysis of children enrolled in Children's Oncology Group AAML0531. Methods: Of 1022 newly diagnosed pediatric patients with de novo AML enrolled on protocol AAML0531, 666 satisfied two criteria for this study: (1) submission of a blood or bone marrow sample for MDF at diagnosis with corresponding CD33 SNP genotype data available, and (2) proper consent for specimen testing. CD33 Expression Levels on Leukemic Blasts The diagnostic AML leukemia population was identified by gating on CD45 versus log-side scatter and CD33 expression levels were determined by measuring the mean fluorescence intensity (MFI) by flow cytometry. For a patient to be considered CD33+ two criteria were required: intensity of CD33 on blasts was at minimum four times the MFI of its correspondent autofluorescent control, and at least 80% of the total blasts were greater than this minimum. Genotyping CD33 SNPs CD33-coding SNP rs12459419-Ala14Val and linked promoter SNP rs3865444 were genotyped using the Sequenome (San Diego, CA) platform at the Biomedical Genomics Center, University of Minnesota. Both SNPs had a call rate of 0.98 and were in Hardy-Weinberg equilibrium (P=0.05). Results: Association of Genotype and cell surface expression of CD33 for AAML0531 patients Of 666 patients, 84% (560/666) were CD33+. CC genotype was observed in 54.5% (305/560) of CD33+ cases, 37.5% (210/560) had CT genotype and 8% (45/560) TT genotype. Of the 16% (106/666) of patients who were CD33 negative, 33% (35/106) had CC genotype, 47% (50/106) CT genotype, and 20% (21/106) had TT genotype. Comprehensively, 340/666 (51%) had CC genotype (51%) and 10% were CD33 negative (35/340). Conversely, out of a total of 66 patients with TT genotype, 45 (68%) were CD33+ and 32% (21/66) were CD33 negative. Conclusions: These results clarify the relationship between the amount of CD33 expressed on AML at diagnosis as measured by MDF and CD33 SNP genotype status. While correlation with clinical outcome analysis is ongoing, these data support inclusion of CD33 SNP genotyping for eligibility of GO therapy. Therefore, the current recommendation for future COG AML clinical protocols is that GO will be administered to patients with CD33 expression as determined by MDF and CC genotype patients regardless of CD33 expression. Disclosures Pardo: Hematologics, Inc: Employment. Eidenschink Brodersen:Hematologics, Inc: Employment. Paine:Hematologics, Inc: Employment. Loken:Hematologics, Inc: Employment, Equity Ownership.

scholarly journals PVRIG is a novel NK cell immune checkpoint receptor in acute myeloid leukemia

Haematologica ◽  
2020 ◽  
pp. 0-0
Author(s):  
Jessica Li ◽  
Sarah Whelan ◽  
Maya F. Kotturi ◽  
Deborah Meyran ◽  
Criselle D’Souza ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Surface ◽  
Nk Cells ◽  
Immune Checkpoint ◽  
Myeloid Leukemia ◽  
Cell Activation ◽  
Nk Cell ◽  
Blocking Antibody ◽  
Poliovirus Receptor ◽  
Acute Myeloid

This study explored the novel immune checkpoint poliovirus receptor-related immunoglobulin domain-containing (PVRIG) in acute myeloid leukemia (AML). We showed that AML patient blasts consistently expressed the PVRIG ligand (poliovirus receptor-related 2, PVRL2). Furthermore, PVRIG blockade significantly enhanced NK cell killing of PVRL2+, poliovirus receptor (PVR)lo AML cell lines, and significantly increased NK cell activation and degranulation in the context of patient primary AML blasts. However, in AML patient bone marrow, NK cell PVRIG expression levels were not increased. To understand how PVRIG blockade might potentially be exploited therapeutically, we investigated the biology of PVRIG and revealed that NK cell activation resulted in reduced PVRIG expression on the cell surface. This occurred whether NK cells were activated by tumour cell recognition, cytokines (IL-2 and IL-12) or activating receptor stimulation (CD16 and NKp46). PVRIG was present at higher levels in the cytoplasm than on the cell surface, particularly on CD56bright NK cells, which further increased cytoplasmic PVRIG levels following IL-2 and IL-12 activation. PVRIG was continually transported to the cell surface via the endoplasmic reticulum (ER) and Golgi in both unstimulated and activated NK cells. Taken together, our findings suggest that anti- PVRIG blocking antibody functions by binding to surface-bound PVRIG, which undergoes rapid turnover in both unstimulated and activated NK cells. We conclude that the PVRIGPVRL2 immune checkpoint axis can feasibly be targeted with PVRIG blocking antibody for NK-mediated immunotherapy of PVRL2+ AML.

Granulocyte Colony-Stimulating Factor (G-CSF) Treatment of Childhood Acute Myeloid Leukemias That Overexpress the Differentiation-Defective G-CSF Receptor Isoform IV Is Associated With a Higher Incidence of Relapse

2010 ◽  
Vol 28 (15) ◽  
pp. 2591-2597 ◽  
Author(s):  
Stephanie Ehlers ◽  
Christin Herbst ◽  
Martin Zimmermann ◽  
Nicole Scharn ◽  
Manuela Germeshausen ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Children And Adolescents ◽  
Cumulative Incidence ◽  
Myeloid Leukemia ◽  
Cell Surface Expression ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Stimulating Factor ◽  
Acute Myeloid ◽  
Factor G

Purpose This prospective, multicenter Acute Myeloid Leukemia Berlin-Frankfurt-Muenster (AML-BFM) 98 study randomly tested the ability of granulocyte colony-stimulating factor (G-CSF) to reduce infectious complications and to improve outcomes in children and adolescents with acute myeloid leukemia (AML). However, a trend toward an increased incidence of relapses in the standard-risk (SR) group after G-CSF treatment was observed. Patients and Methods Of 154 SR patients in the AML-BFM 98 cohort, 50 patients were tested for G-CSF receptor (G-CSFR) RNA isoform I and IV expression, G-CSFR cell surface expression, and acquired mutations in the G-CSFR gene. Results In patients randomly assigned to receive G-CSF after induction, 16 patients overexpressing the G-CSFR isoform IV showed an increased 5-year cumulative incidence of relapse (50% ± 13%) compared with 14 patients with low-level isoform IV expression (14% ± 10%; log-rank P = .04). The level of G-CSFR isoform IV had no significant effect in patients not receiving G-CSF (P = .19). Multivariate analyses of the G-CSF–treated subgroup, including the parameters G-CSFR isoform IV overexpression, sex, and favorable cytogenetics as covariables, revealed the prognostic relevance of G-CSFR isoform IV overexpression for 5-year event-free survival (P = .031) and the 5-year cumulative incidence of relapse (P = .049). Conclusion Our results demonstrate that children and adolescents with AMLs that overexpress the differentiation-defective G-CSFR isoform IV respond to G-CSF administration after induction, but with a significantly higher incidence of relapse.

scholarly journals High-fat diet intensifies MLL-AF9-induced acute myeloid leukemia through activation of the FLT3 signaling in mouse primitive hematopoietic cells

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
François Hermetet ◽  
Rony Mshaik ◽  
John Simonet ◽  
Patrick Callier ◽  
Laurent Delva ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Mouse Model ◽  
Cell Surface ◽  
High Fat Diet ◽  
Myeloid Leukemia ◽  
Potent Inhibitor ◽  
Hematopoietic Cells ◽  
High Fat ◽  
Stat3 Signaling ◽  
Acute Myeloid

Abstract Using a MLL-AF9 knock-in mouse model, we discovered that consumption of a high-fat diet (HFD) accelerates the risk of developing acute myeloid leukemia (AML). This regimen increases the clusterization of FLT3 within lipid rafts on the cell surface of primitive hematopoietic cells, which overactivates this receptor as well as the downstream JAK/STAT signaling known to enhance the transformation of MLL-AF9 knock-in cells. Treatment of mice on a HFD with Quizartinib, a potent inhibitor of FLT3 phosphorylation, inhibits the JAK3/STAT3, signaling and finally antagonizes the accelerated development of AML that occurred following the HFD regimen. We can therefore conclude that, on a mouse model of AML, a HFD enforces the FLT3 signaling pathway on primitive hematopoietic cells and, in turn, improves the oncogenic transformation of MLL-AF9 knock-in cells and the leukemia initiation.

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