OBJECTIVE: To investigate the expressions of CD33 and CD13 in newly diagnosed multiple myeloma (MM) patients and its relationship with prognosis. METHODS: It was retrospectively observed that the expression of CD33 and CD13 in 121 MM patients who were newly diagnosed from January 2014 to January 2020, and the relationship between the expressions of CD33 and CD13 and patients prognosis was analyzed. RESULTS: Among the 121 newly diagnosed MM patients, there were 30 patients (24.8%) in the CD33⁺ group and 12 patients (9.9%) in the CD13⁺ group. Kaplan-Meier analysis showed that, compared with the CD33^- group, the progression-free survival (PFS) time and overall survival (OS) time were significantly shortened in MM patients in CD33⁺ group (PFS 17.5 vs 23 months, P=0.000; OS 18.5 vs 25 months, P=0.000); and the PFS time and OS time of MM patients in the CD13⁺ group were also significantly shortened than those in CD13^- group (PFS 21 vs 22 months, P=0.012; OS 25 vs 26 months, P=0.006). Cox regression analysis showed that CD33 and CD13 were independent adverse prognostic factors in MM patients (CD33: P=0.000;CD13: P=0.003). CONCLUSION CD33 and CD13 are prognostic risk factors in patients with MM.
CD13 Antigens ; Cell Count ; Humans ; Kaplan-Meier Estimate ; Multiple Myeloma ; Prognosis ; Retrospective Studies ; Sialic Acid Binding Ig-like Lectin 3

Objective: To investigate the effect of CD33-targeted bi-specific and tri-specific T-cell engagers on T-cell proliferation and explore their cytotoxicity on leukemia cells. Methods: The CD33-targeted bi-specific T-cell engager (CD33-BiTE) and tri-specific T-cell engager (CD33-TriTE) expression vectors were successfully constructed and expressed through a eukaryotic cell expression system. CD33-BiTE and CD33-TriTE were purified by affinity chromatography. The effects of CD33-BiTE and CD33-TriTE on T cells were analyzed through in vitro experiments. Results: ① CD33-BiTE and CD33-TriTE were successfully constructed and purified and could compete with flow cytometry antibodies for binding to the target cells. ② After 12 days of co-culture with CD33-BiTE and CD33-TriTE, the number of human T cells were expanded to 33.89±19.46 and 81.56±23.62 folds, respectively. CD33-TriTE induced a stronger proliferation of T cells than CD33-BiTE (P<0.05) . ③ Both CD33-BiTE and CD33-TriTE induced specific dose-dependent cytotoxicity on CD33(+) leukemia cells. ④ Compared to CD33-TriTE, leukemia cells were prone to express PD-L1 when co-cultured with T cells and CD33-BiTE. CD33-TriTE induced powerful cytotoxicity on leukemia cells with high PD-L1 expression. Conclusion: CD33-BiTE and CD33-TriTE expression vectors were constructed, and fusion proteins were expressed in eukaryotic cells. Our results support the proliferative and activating effects of BiTE and TriTE on T cells. Compared to that of CD33-BiTE, CD33-TriTE induced a stronger proliferative effect on T cells and a more powerful cytotoxicity on leukemia cells with high PD-L1 expression.
B7-H1 Antigen/pharmacology* ; Humans ; Leukemia, Myeloid, Acute/metabolism* ; Sialic Acid Binding Ig-like Lectin 3/pharmacology* ; T-Lymphocytes

OBJECTIVETo investigate the expression of CD33⁺ HLA-DR⁻ myeloid-derived suppressor cells (MDSCs) in the peripheral blood of patients with renal carcinoma and its correlation with the clinicopathological features of renal cancer.

METHODSForty-four patients with renal carcinoma treated in our hospital between June, 2011 and October, 2012 and 18 healthy volunteers were enrolled in this study. Flow cytometry was performed to detect CD33⁺ HLA-DR⁻ MDSCs in the peripheral blood, and its correlation with the clinicopathological features of the patients were analyzed.

RESULTSThe positivity rate of CD33⁺ HLA-DR⁻ MDSCs in the peripheral blood was significantly higher in the cancer patients than in the healthy controls [(1.91 ± 0.66)% vs (0.62 ± 0.22)%, P<0.001]. The expression levels of CD33⁺ HLA-DR⁻ MDSCs in patients with renal carcinoma showed significant differences between stage I+II [(1.46 ± 0.44)%] and stage III [(2.04 ± 0.35)%] patients (P<0.01) and between stage III and stage IV patients [(2.50 ± 0.64)%] (P<0.05), but did not differ significantly in respect of age or gender.

CONCLUSIONCD33⁺ HLA-DR⁻ MDSCs expression in the peripheral blood is associated with tumor stage and differentiation in renal carcinoma and may play an important role in predicting the prognosis and tumor immunology of renal carcinoma.

Adult ; Aged ; Case-Control Studies ; Female ; Flow Cytometry ; HLA-DR Antigens ; metabolism ; Humans ; Immunophenotyping ; Kidney Neoplasms ; blood ; immunology ; Male ; Middle Aged ; Myeloid Cells ; cytology ; metabolism ; Neoplasm Staging ; Prognosis ; Sialic Acid Binding Ig-like Lectin 3 ; metabolism ; Young Adult

This study was aimed to investigate clinical and prognostic significances of 4 target antigens (CD19, CD20, CD22 and CD33) for antibody-based immunotherapy and to evaluate the applications of these antibody-based target therapy to adult acute lymphoblastic leukemia (ALL). The immunophenotype of 220 adult patients with ALL were analyzed by four-color flow Cytometry, and cytogenetic and molecular parameters were detected by conventional cytogenetics, fluorescence in situ hybridization, real-time quantitative PCR, nested PCR and DNA sequencing. The results showed that CD19 positive (CD19(+)) cases were more in female (46.4% vs. 23.4%, P = 0.006), elderly patients aged > 60 years (14.4% vs. 2.1%, P = 0.022), CD33(+) co-expression cases (47.8% vs. 12.0%, P = 0.001) and genetic high-risk group (55.8% vs. 20.8%, P = 0.002) compared with CD19 negative (CD19(-)) cases; CD20(+) cases had lower co-expression of CD13 than CD20(-) cases (31.6% vs.67.1%, P = 0.000) and no significant prognostic indications for CD20(+) was observed; CD22(+) cases had higher relapse rate at 12-month than CD22(-) cases (93.9% vs.57.1%, P = 0.041) in B-ALL patients; CD33(+) cases had higher incidence of Ph(+) than CD33(-) cases (43.5% vs.19.4%, P = 0.007) and significantly correlated with Ph(+) (r = 0.261, P = 0.006). It is concluded that elucidation of the characteristics of the target antigens (CD19, CD20, CD22, CD33) used for antibody-based immunotherapy will help hematologists making the correct decision whether and when to use these antibody-based target therapies.
Adolescent ; Adult ; Aged ; Antigens, CD19 ; immunology ; Antigens, CD20 ; immunology ; Child ; Female ; Humans ; Immunophenotyping ; Male ; Middle Aged ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; immunology ; Sialic Acid Binding Ig-like Lectin 2 ; immunology ; Sialic Acid Binding Ig-like Lectin 3 ; immunology ; Young Adult

This study is to investigate the effect of downregulation histone deacetylases 1 (HDAC1) gene by the technology of RNA interference on the differentiation of HL-60 cells line. The optimal segment targeting HDAC1 gene was designed and transfected into HL-60 cells by Lipofectamine 2000. The HDAC1 mRNA and protein level were detected by RT-PCR and Western blotting. The morphologic change of HL-60 cells was detected by an optical microscope with Wright-Giemsa. Cell differentiation was tested by NBT reduction assay. Expression of CD13, CD33 and CD14 was measured by flow cytometry. The results indicated that HDAC1 mRNA and protein were markedly suppressed by the siRNA targeting HDAC1 in a concentration-dependent manner. HDAC1 siRNA promoted cell differentiation. HL-60 cells became more mature in morphology after transfected to HDAC1 siRNA at a concentration of 30-60 nmol x L(-1) for 24 h. NBT reduction ability of HDAC1 siRNA with 30 nmol x L(-1) was 0.31 +/- 0.09, compared with negative control (0.20 +/- 0.02) (t = -3.1, P < 0.01), and with 60 nmol x L(-1) was 0.25 +/- 0.02 in comparison with negative control (0.21 +/- 0.04) (t = -2.12, P < 0.05). But it has no change in HDAC1 siRNA > or = 120 nmol x L(-1). After transfection with 60 nmol x L(-1) HDAC1 siRNA to HL-60 cells, the expression of CD13 was (96.50 +/- 0.70)% in compared to siRNA-NC (3.39 +/- 0.68) % (t = 164.9, P < 0.000 5), CD33 was (66.73 +/- 0.50) % in compared to siRNA-NC (96.80 +/- 1.70) % (t = 43.4, P < 0.000 5). CD14 was (0.53 +/- 0.00) % by comparison with siRNA-NC (0.49 +/- 0.02) % (t = -0.97, P > 0.1). HDAC1 siRNA promoted cell differentiation in indicated concentration. HDAC1 might be one of the targets of gene therapy for leukemia.
CD13 Antigens ; metabolism ; Cell Differentiation ; Down-Regulation ; HL-60 Cells ; Histone Deacetylase 1 ; genetics ; metabolism ; Humans ; Lipopolysaccharide Receptors ; metabolism ; RNA Interference ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; genetics ; Sialic Acid Binding Ig-like Lectin 3 ; metabolism ; Transfection

OBJECTIVE: To screen aptamers binding CD33+/CD34- cells from patients with acute myeloblastic leukemia M2 subtype (AML-M2). METHODS: CD33+/CD34- cells from patients with AML-M2 were taken as targeted cells, CD33+/ CD34- cells from normal people were taken as anti-selecting cells, and aptamers in the single strand deoxyribonucleic acid (ssDNA) library were then selected repeatedly by cell-systematic evolution of ligands by exponential enrichment (C-SELEX) technology, and amplified by polymerase chain reaction (PCR) to generate sub-ssDNA library. During the experiment, PCR amplification with fluorescently labeled primer and flow cytometry were performed to analyze the aptamers'enrichment of sub-library, and the final round product of the sub-ssDNA library was cloned. After the sequencing, the primary and secondary structures of the aptamers were analyzed. RESULTS: Electrophoresis indicated that the product of PCR amplification for each round subssDNA library was able to see a clear DNA band in the agarose gel. After 13 rounds of screening, the fluorescence intensity of the sub-ssDNA library binding the cells ranged from 2.14% to 51.12%, reaching a steady state at the 13th round. A total of 30 clones were selected and sequenced, 22 of which contained 1 of the 4 conserved sequences of AAGTA, TATCT, AGATG and AAATT in their primary structure, but the remained eight aptamers contained none of the conserved sequence. Secondary structure analysis indicated that four stem-loops and loop simulation convex structures existed in the aptamers. CONCLUSION C-SELEX technology can be used to screen the aptamers binding primary cells from patients with leukemia. The aptamers selected from the CD33+/CD34- cells from the patients of AML-M2 subtype might be used for the diagnosis and treatment for leukemia.
Adolescent ; Adult ; Antigens, CD34 ; genetics ; immunology ; Antigens, Differentiation, Myelomonocytic ; genetics ; immunology ; Aptamers, Nucleotide ; genetics ; metabolism ; DNA, Single-Stranded ; genetics ; Female ; Humans ; Leukemia, Myeloid, Acute ; genetics ; immunology ; Male ; Middle Aged ; SELEX Aptamer Technique ; Sialic Acid Binding Ig-like Lectin 3 ; genetics ; immunology ; Young Adult

The objective of this study was to investigate the immunophenotype of T-lineage acute lymphoid leukemia (T-ALL) and to find valuable immunologic markers in T-ALL diagnosis and therapy. Four-color multiparametric flow cytometry(FCM) with CD45/SSC gating was used for immunophenotyping of 95 patients with newly diagnosed T-ALL. The results demonstrated that T-ALL occurred more frequently in males younger than 30 years of age and was usually accompanied by a high WBC count and tumor mass at diagnosis. Univariate analysis showed an influence on achievement of CR1 for age (< 30 years) but not for WBC count and tumor mass. According to WHO (2008) classification of tumors of haematopoietic and lymphoid tissues, 87 patients with confirmed subtype included 27 cases of Pro-T-ALL (31.0%), 31 cases of Pre-T-ALL (35.6%), 23 cases of cortical-T-ALL (26.4%), 6 cases of medullary-T-ALL (6.9%). CD34 expression in Pro-T-ALL was significantly higher than that of Pre-T-ALL (p = 0.001). After the first chemotherapy, the complete remission rate in Pro-T-ALL was statistically lower than that of Pre-T-ALL. Besides, the complete remission rate of immature T-ALL (including Pro-T-ALL and Pre-T-ALL) was also significantly lower than that in mature T-ALL (including cortical-T-ALL and medullary-T-ALL). Myeloid antigen (CD13, CD33) expression was associated with T-ALL subtype and treatment effect. While 66.7% of CD13(+) patients belonged to Pre-T-ALL, most (60.0%) of CD33(+) patients were classified into Pro-T-ALL; CD13 expression had no effect on CR1 rate whereas CD33(+) patients had worse treatment effect compared with CD33(-) groups (p = 0.001). Notably, the expression of CD117 reached up to 26.7% and the positive cases were primarily distributed in pro-T-TAll and pre-T-ALL. It is found that CD117 expression in CD34(-) group was homogeneous and CD117 expression level was less than 10% in 73.2% patients, but CD117 expression level in CD34(+) group was not homogenous, in which group the CD117 expression levels < 10%, 10% - 20% and > 20% were 44.2%, 17.3% and 38.5% respectively. As compared with CD34(-) group, the proportion of patients with CD117 expression levels < 10%, > 20% in CD34(+) group was higher, and there was significant difference between these 2 group. It is concluded that immunophenotype has great value in T-ALL diagnosis, classification as well as treatment. Flow cytometry provides access to find valuable immunologic markers for T-ALL biological research.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; CD13 Antigens ; metabolism ; Child ; Child, Preschool ; Female ; Humans ; Immunophenotyping ; Infant ; Male ; Middle Aged ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma ; classification ; immunology ; therapy ; Proto-Oncogene Proteins c-kit ; metabolism ; Sialic Acid Binding Ig-like Lectin 3 ; metabolism ; Young Adult

A little is known about the specific marker on the surface of acute leukemia cells, leading to the lack of the specific diagnosis method for acute leukemia. Therefore, in this study, cell-systematic evolution of ligands by exponential enrichment (cSELEX) was performed to screen the aptamers binding to CD33(+)/CD34(+) cells from the patients with acute myeloblastic leukemia (AML) of M(2) subtype (AML-M₂) so as to provide the basis for finding the specific marker on the surface of AML-M(2) CD33(+)/CD34(+) cells. Firstly, AML-M₂ CD33(+)/CD34(+) cells were sorted and used as targeted cells, and normal CD33(+)/CD34(+)cells were used as counter-targeted cells; the aptamers binding to CD33(+)/CD34(+) cells from patients with AML-M₂ were screened from the single strand deoxyribonucleic acid (ssDNA) library by cSELEX. Subsequently, each aptamer structure was analyzed after cloning and sequencing. The results indicated that after 13 round of screenings, the enrichment of aptamers in the ssDNA library was ranged from 0.7% to 52.9%, and reached steady state at 13th round screening. Sequence analysis for 30 aptamers showed that most of the aptamers born one of the three conserved sequences of CCCCT, CTCTC, and CTCAC. Secondary structure analysis indicated that three different secondary structures existed in these aptamers. It is concluded that the aptamers binding to the AML-M(2) CD33(+)/CD34(+) cells are successfully screened, which lay the basis for further looking for the specific marker on the surface of AML-M₂ CD33(+)/CD34(+) cells, and the molecular diagnosis of the AML-M₂ leukemia.
Antigens, CD ; genetics ; immunology ; Antigens, CD34 ; genetics ; immunology ; Antigens, Differentiation, Myelomonocytic ; genetics ; immunology ; Aptamers, Nucleotide ; metabolism ; Biomarkers ; Flow Cytometry ; Humans ; Immunophenotyping ; Leukemia, Myeloid, Acute ; genetics ; immunology ; Nucleic Acid Conformation ; SELEX Aptamer Technique ; Sialic Acid Binding Ig-like Lectin 3

OBJECTIVETo isolate CD133(+)/CD44(+)/ESA(+) subsets cells from SW480 colon cancer cells, and to observe the tumor formation.

METHODCD133(+)/CD44(+)/ESA(+) subsets cells, CD133(-)/CD44(+)/ESA(+) subsets cells and CD133(-)/CD44(-)/ESA(-) subsets cell were sorted by flow cytometry from SW480 colon cancer cells, then three subsets were separately inoculated in five NOD/SCID mice and the growth rates were calculated.

RESULTThe proportion of CD133(-)/CD44(-)/ESA(-), CD133(-)/CD44(+)/ESA(+) and CD133(+)/CD44(+)/ESA(+) subsets cells in SW480 cells were (86.38±10.23)%,(1.26±0.28)% and(0.38±0.07)%. After inoculation, tumor nodules could be formed three days later in CD133(+)/CD44(+)/ESA(+) group, and they could be formed 9 days later in CD133(-)/CD44(+)/ESA(+) group, while they could be formed 15 days later in CD133(-)/CD44(-)/ESA(-) group. Eighteen days later, tumor sizes in three groups were(13.82±5.04) mm(3), (9.25±4.57) mm(3) and (4.76±3.92) mm(3) respectively, and the differences were statistically significant(P<0.05).

CONCLUSIONESA(+)-CD44(+) is one of the surface markers for colonic cancer stem cells, and CD133(+)-CD44(+)-ESA(+) cells are SW480-like cancer stem cells.

Animals ; Biomarkers, Tumor ; Cell Line, Tumor ; Colonic Neoplasms ; pathology ; Flow Cytometry ; Humans ; Hyaluronan Receptors ; Mice ; Mice, Inbred NOD ; Mice, SCID ; Neoplastic Stem Cells ; cytology ; Sialic Acid Binding Ig-like Lectin 3

The aim of study was to investigate the immunophenotype characteristics and prognosis of acute leukemia patients with cross-expressing lymphoid and myeloid lineage-associated antigens. The immunophenotypes of leukemic cells were examined by using flow cytometry. All patients were classified into several groups according to FAB subtypes and immunophenotyping. The cross-expressed antigens analyzed for AML included CD2, CD7, CD19, CD56 and other co-expressed lymphoid antigens. The myeloid antigens analyzed for ALL included CD13 and co-expressed CD13/CD33. ALL and AML patients without expression of cross-expressing antigens were selected as control. Complete remission (CR) ratio and relapse-free survival (RFS) of patients in all groups were compared. The results indicated that among 161 patients analyzed, 91 cases of AML with cross-expressing lymphoid and myeloid antigens included that 24 cases of AML expressed lymphoid surface marker-CD7, namely CD7(+) AML, 14 cases of AML only expressed lymphoid surface marker-CD19, namely CD19(+) AML, 8 cases of AML expressed lymphoid surface marker-CD2 (including CD2/CD19 co-expressed), namely CD2(+) AML, 10 cases of AML expressed lymphoid surface marker-CD56 (including CD56/CD19 or CD56/CD2 co-expressed), namely CD56(+) AML, 16 cases of AML expressed two or more lymphoid surface markers, namely Ly ≥ 2(+) AML, 9 cases of ALL expressed myeloid surface markers CD13, namely CD13(+) ALL, 10 cases of ALL expressed myeloid surface markers CD13 and CD33, namely CD13/CD33(+) ALL. 29 cases of ALL did not expressed myeloid surface markers, namely My(-) ALL, and 41 case of AML did not expressed lymphoid surface markers, namely Ly(-) AML. CR ratio and RFS of Ly ≥ 2(+) AML patients were lower than those of Ly(-) AML patients. RFS of CD56(+) AML patients was lower, but CR ratio had no significant difference, when compared with Ly(-) AML patients. CR ratio and RFS of other AML patients with cross-expressing antigens had no significant difference when compared with Ly(-) AML patients. CR ratio and RFS of CD13(+) ALL and CD13/CD33(+) ALL patients had no significant difference when compared with My(-) ALL patients. It is concluded that the importance of cross-expressing antigens for prognosis of patients should be analyzed concretely. CD56(+) AML and Ly ≥ 2(+) AML have bad prognosis, while other cross-expressed lymphoid and myeloid lineage-associated antigens have no impact on prognosis of acute leukemia patients.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antigens, CD ; immunology ; Antigens, Differentiation, Myelomonocytic ; immunology ; CD13 Antigens ; immunology ; CD56 Antigen ; immunology ; Child ; Child, Preschool ; Female ; Humans ; Immunophenotyping ; Infant ; Leukemia, Myeloid, Acute ; classification ; immunology ; Male ; Middle Aged ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; classification ; immunology ; Prognosis ; Sialic Acid Binding Ig-like Lectin 3 ; Young Adult