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Journal of Translational Medicine Mar 2024Targeting CD47/SIRPα axis has emerged as a promising strategy in cancer immunotherapy. Despite the encouraging clinical efficacy observed in hematologic malignancies...
BACKGROUND
Targeting CD47/SIRPα axis has emerged as a promising strategy in cancer immunotherapy. Despite the encouraging clinical efficacy observed in hematologic malignancies through CD47-SIRPα blockade, there are safety concerns related to the binding of anti-CD47 antibodies to CD47 on the membrane of peripheral blood cells.
METHODS
In order to enhance the selectivity and therapeutic efficacy of the antibody, we developed a humanized anti-CD47 monoclonal antibody called Gentulizumab (GenSci059). The binding capacity of GenSci059 to CD47 was evaluated using flow cytometry and surface plasmon resonance (SPR) methods, the inhibitory effect of GenSci059 on the CD47-SIRPα interaction was evaluated through competitive ELISA assays. The anti-tumor activity of GenSci059 was assessed using in vitro macrophage models and in vivo patient-derived xenograft (PDX) models. To evaluate the safety profile of GenSci059, binding assays were conducted using blood cells. Additionally, we investigated the underlying mechanisms contributing to the weaker binding of GenSci059 to erythrocytes. Finally, toxicity studies were performed in non-human primates to assess the potential risks associated with GenSci059.
RESULTS
GenSci059 displayed strong binding to CD47 in both human and monkey, and effectively inhibited the CD47-SIRPα interaction. With doses ranging from 5 to 20 mg/kg, GenSci059 demonstrated potent inhibition of the growth of subcutaneous tumor with the inhibition rates ranged from 30.3% to complete regression. Combination of GenSci059 with 2.5 mg/kg Rituximab at a dose of 2.5 mg/kg showed enhanced tumor inhibition compared to monotherapy, exhibiting synergistic effects. GenSci059 exhibited minimal binding to hRBCs compared to Hu5F9-G4. The binding of GenSci059 to CD47 depended on the cyclization of N-terminal pyroglutamic acid and the spatial conformation of CD47, but was not affected by its glycosylation modifications. A maximum tolerated dose (MTD) of 450 mg/kg was observed for GenSci059, and no significant adverse effects were observed in repeated dosages up to 10 + 300 mg/kg, indicating a favorable safety profile.
CONCLUSION
GenSci059 selectively binds to CD47, effectively blocks the CD47/SIRPα axis signaling pathway and enhances the phagocytosis effects of macrophages toward tumor cells. This monoclonal antibody demonstrates potent antitumor activity and exhibits a favorable safety profile, positioning it as a promising and effective therapeutic option for cancer.
Topics: Animals; Humans; CD47 Antigen; Neoplasms; Phagocytosis; Macrophages; Antibodies, Monoclonal; Immunotherapy; Disease Models, Animal; Antigens, Differentiation
PubMed: 38429732
DOI: 10.1186/s12967-023-04710-6 -
Immunity Sep 1997
Review
Topics: ADP-ribosyl Cyclase; ADP-ribosyl Cyclase 1; Animals; Antigens, CD; Antigens, Differentiation; Biomarkers; Cell Differentiation; Humans; Lymphocytes; Membrane Glycoproteins; Mice; NAD+ Nucleosidase; Signal Transduction
PubMed: 9324352
DOI: 10.1016/s1074-7613(00)80353-2 -
Journal of Neuroimmunology 1990Antigens shared by the immune and central nervous systems (CNS) have been described repeatedly. The present study reports the expression of the CD6 lymphocyte...
Antigens shared by the immune and central nervous systems (CNS) have been described repeatedly. The present study reports the expression of the CD6 lymphocyte differentiation antigen in normal human brain evidenced by immunohistochemistry and Northern blot analysis. A panel of various anti-CD6 monoclonal antibodies (mabs) tested on serial cryostat sections identified CD6-positive cells randomly scattered in parenchyma of all examined brain areas. Northern blot analysis with a highly sensitive cRNA probe revealed a 3.1 kb CD6-specific mRNA in various brain regions, especially in basal ganglia and cortex cerebellum. Staining with mabs raised against different hematopoietic cell types, as well as hybridization with probes specific for the beta- and gamma-T cell receptor (TCR) chains support the notion that CD6 is expressed by original brain cells. The nature of the CD6-positive cell type and possible functions of shared antigens in immune and nervous systems are discussed.
Topics: Antibodies, Monoclonal; Antigens, CD; Antigens, Differentiation, T-Lymphocyte; Blotting, Northern; Brain; Humans; Immunohistochemistry; RNA, Messenger; Receptors, Antigen, T-Cell; T-Lymphocytes
PubMed: 2211985
DOI: 10.1016/0165-5728(90)90162-g -
Blood Aug 1995Leukemia may be viewed as a clonal expansion of blast cells; however, the role of primitive cells and/or stem cells in disease etiology and progression is unclear. We...
Leukemia may be viewed as a clonal expansion of blast cells; however, the role of primitive cells and/or stem cells in disease etiology and progression is unclear. We investigated stem cell involvement in leukemia using fluorescence in situ hybridization (FISH), immunofluorescence labeling of hematopoietic subpopulations, and flow cytometric analysis/sorting to discriminate and quantify cytogenetically aberrant stem cells in 12 acute myeloid leukemia (AML) and three myelodysplastic (MDS) specimens. Flow cytometric analysis and sorting were used to discriminate and collect a primitive subpopulation enriched in stem cells expressing CD34+ and lacking CD33 and CD38 (CD34+lin-). A subpopulation containing progenitors and differentiating myeloid cells expressed CD34, CD33, and CD38 (CD34+lin+). Nine specimens contained less than 10% CD34+ cells and, thus, were considered to be CD34- leukemias. Mature lymphoid, myeloid, and erythroid subpopulations were sorted on the basis of antigen-linked immunofluorescence. Cytogenetically aberrant cells in sorted subpopulations were identified using FISH with enumerator probes selected on the basis of diagnosis karyotype. Cytogenetically aberrant CD34+lin- cells were present at frequencies between 9% and 99% in all specimens. CD34+lin- cytogenetically aberrant cells comprised between 0.05% and 11.9% of the marrow/blood specimens. Cytogenetically aberrant CD34+lin+ cells constituted 0.01% tp 56% of the marrow/blood population. These data demonstrate that aberrant cells are present in primitive CD34+ stem cell compartments, even in CD34- leukemias. Stem cell involvement was confirmed further by sorting lymphoid and erythroid subpopulations from eight specimens in which the predominant leukemic population lacked lymphoid/erythroid differentiation markers. In these specimens, as well as in multiple lineages, suggests involvement of a cell(s) with multilineage capabilities. The ability of aberrant CD34+lin- stem cells to contribute to clonal and compartment expansion within immunofluorescently defined subpopulations was evaluated to explore the functional phenotype of aberrant CD34+lin- cells. Analysis of compartment size and aberrant cell frequency suggests that frequency of cytogenetically aberrant stem cells is uncoupled from compartment size. These data suggest that cytogenetically aberrant cells in the primitive compartment show varying abilities to expand primitive compartments. Cytogenetically aberrant CD34+lin- cells precede the blast subpopulation in hierarchical maturation and may in some cases by considered preleukemic, requiring maturation or additional mutations before transformation (eg, compartmental expansion) occurs.
Topics: ADP-ribosyl Cyclase; ADP-ribosyl Cyclase 1; Acute Disease; Adult; Aged; Antigens, CD; Antigens, CD34; Antigens, Differentiation; Antigens, Differentiation, Myelomonocytic; Chromosome Aberrations; Chromosome Disorders; Female; Humans; Immunophenotyping; In Situ Hybridization, Fluorescence; Leukemia, Myeloid; Male; Membrane Glycoproteins; Middle Aged; Myelodysplastic Syndromes; N-Glycosyl Hydrolases; Neoplastic Stem Cells; Sialic Acid Binding Ig-like Lectin 3
PubMed: 7542497
DOI: No ID Found -
Scientific Reports Mar 2019Numerous in vitro models endeavour to mimic the characteristics of primary human hepatocytes for applications in regenerative medicine and pharmaceutical science....
Numerous in vitro models endeavour to mimic the characteristics of primary human hepatocytes for applications in regenerative medicine and pharmaceutical science. Mature hepatocyte-like cells (HLCs) derived from human induced pluripotent stem cells (hiPSCs) are one such in vitro model. Due to insufficiencies in transcriptome to proteome correlation, characterising the proteome of HLCs is essential to provide a suitable framework for their continual optimization. Here we interrogated the proteome during stepwise differentiation of hiPSCs into HLCs over 40 days. Whole cell protein lysates were collected and analysed using stabled isotope labelled mass spectrometry based proteomics. Quantitative proteomics identified over 6,000 proteins in duplicate multiplexed labelling experiments across two different time course series. Inductive cues in differentiation promoted sequential acquisition of hepatocyte specific markers. Analysis of proteins classically assigned as hepatic markers demonstrated trends towards maximum relative abundance between differentiation day 30 and 32. Characterisation of abundant proteins in whole cells provided evidence of the time dependent transition towards proteins corresponding with the functional repertoire of the liver. This data highlights how far the proteome of undifferentiated precursors have progressed to acquire a hepatic phenotype and constructs a platform for optimisation and improved maturation of HLC differentiation.
Topics: Antigens, Differentiation; Cell Differentiation; Hepatocytes; Humans; Induced Pluripotent Stem Cells; Proteomics
PubMed: 30824743
DOI: 10.1038/s41598-019-39400-1 -
Archives of Pathology & Laboratory... Jan 2003Acute leukemia displays characteristic patterns of surface antigen expression (CD antigens), which facilitate their identification and proper classification and hence...
CONTEXT
Acute leukemia displays characteristic patterns of surface antigen expression (CD antigens), which facilitate their identification and proper classification and hence play an important role in instituting proper treatment plans. In addition to enzyme cytochemical analysis, multiparameter flow cytometric analysis has become commonplace in most laboratories for that purpose. The essential role and caveats of flow cytometry in that regard, however, have received little scrutiny.
OBJECTIVE
To evaluate the expression of commonly used immunomarkers and patterns in various acute leukemias to help define the best use and role of multiparameter flow cytometry in the diagnosis and proper classification of acute leukemias.
DESIGN
We have retrospectively analyzed the immunophenotypic data from 508 de novo adult and pediatric acute leukemia patients, as studied using multiparameter flow cytometry in addition to routine morphologic and enzyme cytochemical analysis. Cytogenetic and/or molecular data were correlated in all 41 cases of acute promyelocytic leukemia (APL) and in 203 other cases of acute leukemia where those data were available. We have also determined the positive and negative predictive values of a combined CD34 and HLA-DR expression pattern for the differentiation of APL from other myeloid leukemias.
RESULTS
In acute myeloid leukemia (AML) other than APL, expression of CD34 was seen in 62% and expression of HLA-DR in 86% of all cases. Twenty-six (10%) of 259 cases of non-APL AML were negative for both CD34 and HLA-DR as opposed to 33 (80%) of 41 cases of APL. None of the cases of APL were positive for both CD34 and HLA-DR in contrast to 149 (58%) of 259 cases of non-APL AML. Fifty-three cases were found to be examples of minimally differentiated AML (AML M0) based on the lack of expression of cytoplasmic CD3 and cytoplasmic CD79a and expression of one or more myelomonocytic-associated antigens and/or myeloperoxidase. Expression of CD20 was seen in 40 (24%) of 168 cases of precursor B-cell acute lymphoblastic leukemia (pB-ALL) and 52 (29%) lacked CD34 expression. Five of 180 cases of pB-ALL and 2 cases of precursor T-cell ALL (pT-ALL) were negative for terminal deoxynucleotidyl transferase (TdT). Aside from cytoplasmic CD3, CD5 and CD7 were the most sensitive antigens present in all 21 cases of pT-ALL. CD33 was more sensitive but less specific than CD13 for myeloid lineage.
CONCLUSION
Aside from identification of blasts, flow cytometry was found to be especially useful in the correct identification of AML M0, differentiation of APL from AML M1/M2, and correct identification of TdT-negative ALL and unusual variants, such as transitional B-cell ALL and undifferentiated and biphenotypic acute leukemias.
Topics: Acute Disease; Adult; Antigens, CD34; Antigens, Differentiation, B-Lymphocyte; Antigens, Differentiation, Myelomonocytic; Antigens, Differentiation, T-Lymphocyte; Cytogenetic Analysis; DNA Nucleotidylexotransferase; Flow Cytometry; HLA-DR Antigens; Humans; Immunophenotyping; Killer Cells, Natural; Leukemia; Megakaryocytes; Peroxidase; Proto-Oncogene Proteins c-kit; Retrospective Studies
PubMed: 12521365
DOI: 10.5858/2003-127-42-FCAOA -
Journal of Molecular Biology Jan 2008Sialic acid (Sia) Ig-like binding lectins are important mediators of recognition and signaling events among myeloid cells. To investigate the molecular mechanism...
Sialic acid (Sia) Ig-like binding lectins are important mediators of recognition and signaling events among myeloid cells. To investigate the molecular mechanism underlying sialic acid Ig-like lectin (Siglec) functions, we determined the crystal structure of the two N-terminal extracellular domains of human myeloid cell inhibitory receptor Siglec-5 (CD170) and its complexes with two sialylated carbohydrates. The native structure revealed an unusual conformation of the CC' ligand specificity loop and a unique interdomain disulfide bond. The alpha(2,3)- and alpha(2,6)-sialyllactose complexed structures showed a conserved Sia recognition motif that involves both Arg124 and a portion of the G-strand in the V-set domain forming beta-sheet-like hydrogen bonds with the glycerol side chain of the Sia. Only few protein contacts to the subterminal sugars are observed and mediated by the highly variable GG' linker and CC' loop. These structural observations, in conjunction with surface plasmon resonance binding assays, provide mechanistic insights into linkage-dependent Siglec carbohydrate recognition and suggest that Siglec-5 and other CD33-related Siglec receptors are more promiscuous in sialoglycan recognition than previously understood.
Topics: Amino Acid Sequence; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Binding Sites; Crystallization; Disulfides; Escherichia coli; Humans; Hydrogen Bonding; Kinetics; Lectins; Ligands; Models, Chemical; Models, Molecular; Molecular Sequence Data; Polysaccharides; Protein Binding; Protein Conformation; Protein Structure, Secondary; Protein Structure, Tertiary; Recombinant Proteins; Sequence Homology, Amino Acid; Sialic Acid Binding Immunoglobulin-like Lectins; Static Electricity; Surface Plasmon Resonance; X-Ray Diffraction
PubMed: 18022638
DOI: 10.1016/j.jmb.2007.10.009 -
Journal of Cellular and Molecular... 2003NK cells express receptors characterized by opposite functions that finely regulate their activities. Among inhibitory receptors, some are specific for different groups... (Review)
Review
NK cells express receptors characterized by opposite functions that finely regulate their activities. Among inhibitory receptors, some are specific for different groups of MHC class I alleles, while others are still orphan receptors. On the contrary, various activating receptors are involved in the triggering of NK-mediated natural cytotoxicity. In general, their engagement induces human NK cells to kill target cells that are either HLA class I-negative or -deficient. Thus, the process of NK cell triggering mediated by Natural Cytotoxicity Receptors can be mainly considered as a non MHC-restricted mechanism. Here, a brief description of the molecular nature of these receptors, as well as, of their 3D-structures and of the implications for ligand recognition, is given.
Topics: Antigens, CD; Antigens, Differentiation, Myelomonocytic; Chromosomes, Human, Pair 17; Cytotoxicity, Immunologic; Humans; Killer Cells, Natural; Lectins; Models, Immunological; Models, Molecular; Multigene Family; Natural Cytotoxicity Triggering Receptor 1; Natural Cytotoxicity Triggering Receptor 2; Psoriasis; Receptors, Immunologic; Receptors, KIR; Receptors, Natural Killer Cell
PubMed: 14754506
DOI: 10.1111/j.1582-4934.2003.tb00240.x -
Immunology Today Oct 1994Among vascular cell adhesion molecules, platelet-endothelial cell adhesion molecule (PECAM-1/CD31) has the distinctive feature of being expressed on several of the major... (Review)
Review
Among vascular cell adhesion molecules, platelet-endothelial cell adhesion molecule (PECAM-1/CD31) has the distinctive feature of being expressed on several of the major cell types associated with the vascular compartment. This makes it uniquely positioned to mediate multiple and important cell-cell interactions involving platelets, leukocytes and endothelial cells. Thus, PECAM-1 may represent a potential target for new therapeutic agents directed at a variety of pathological states.
Topics: Alternative Splicing; Animals; Antibodies, Monoclonal; Antigens, Differentiation, Myelomonocytic; Cell Adhesion; Cell Adhesion Molecules; Fluorescent Antibody Technique; Humans; Platelet Endothelial Cell Adhesion Molecule-1
PubMed: 7945775
DOI: 10.1016/0167-5699(94)90195-3 -
Blood Jul 1992Twenty-three acute myelocytic leukemia (AML) patients with t(8;21) chromosomal abnormality, all classified as M2 (French-American-British [FAB] classification), were...
Phenotypical characteristics of acute myelocytic leukemia associated with the t(8;21)(q22;q22) chromosomal abnormality: frequent expression of immature B-cell antigen CD19 together with stem cell antigen CD34.
Twenty-three acute myelocytic leukemia (AML) patients with t(8;21) chromosomal abnormality, all classified as M2 (French-American-British [FAB] classification), were investigated. Blastic cells from all patients were positive for the stem cell-associated antigens, CD34 and HLA-DR, and the immature myeloid antigens, CD13 and CD33. The nonblastic leukemic cells expressed the more mature myeloid antigens, CD11b and CD15, with loss of the immature phenotype. The incidence of positivities for the stem cell-associated antigens, CD34 and HLA-DR, in t(8;21) AML cells was significantly higher in comparison with those in other AML showing granulocytic differentiation (M2 or M3). AML cells with t(8;21) also showed some phenotypic abnormalities. Frequent expression of CD19 was found in the blastic population of t(8;21) AML (18 of 23 cases) without other B-cell antigens and Ig gene rearrangements. CD19 expression was confirmed by immunocytochemistry and Northern blotting. The CD19+ blastic cells coexpressed both CD34 and HLA-DR. In addition, CD33+ cells among the blastic fraction in t(8;21) AML cells were fewer in number than in those of M2 or M3 AML without t(8;21). Our findings indicate that leukemic blasts of t(8;21) AML commonly express CD19 while preserving the stem cell-associated antigens, and differentiate into the granulocytic pathway with discordant maturation such as low CD33 expression.
Topics: Adult; Antigens, CD; Antigens, CD19; Antigens, CD20; Antigens, CD34; Antigens, Differentiation, B-Lymphocyte; B-Lymphocytes; Base Sequence; Chromosome Aberrations; Chromosome Disorders; Chromosomes, Human, Pair 21; Chromosomes, Human, Pair 8; Humans; Karyotyping; Leukemia, Myeloid, Acute; Molecular Sequence Data; Oligodeoxyribonucleotides; Phenotype; Polymerase Chain Reaction; Stem Cells; Translocation, Genetic
PubMed: 1378322
DOI: No ID Found