The study aimed to construct the second and third generation chimeric antigen receptor T cells (CAR-T) targeting the c-mesenchymal-epithelial transition factor (c-Met) protein, and observe their killing effect on human serous ovarian cancer cell SKOV-3. The expression of MET gene in ovarian serous cystadenocarcinoma, the correlation between MET gene expression and the abundance of immune cell infiltration, and the effect of MET gene expression on the tissue function of ovarian serous cystadenocarcinoma were analyzed by bioinformatics. The expression of c-Met in ovarian cancer tissues and adjacent tissues was detected by immunohistochemical staining. The second and third generation c-Met CAR-T cells, namely c-Met CAR-T(2G/3G), were prepared by lentivirus infection, and the cell subsets and infection efficiency were detected by flow cytometry. Using CD19 CAR-T and activated T cells as control groups and A2780 cells with c-Met negative expression as Non target groups, the kill efficiency on SKOV-3 cells with c-Met positive expression, cytokine release and cell proliferation of c-Met CAR-T(2G/3G) were explored by lactate dehydrogenase (LDH) release, ELISA and CCK-8 respectively. The results showed that MET gene expression was significantly up-regulated in ovarian cancer tissues compared with normal tissues, which was consistent with the immunohistochemistry results. However, in all pathological stages, there was no obvious difference in MET expression and no correlation between MET gene expression and the race and age of ovarian cancer patients. The second generation and third generation c-Met CAR-T cells were successfully constructed. After lentivirus infection, the proportion of CD8⁺ T cells in c-Met CAR-T(2G) was upregulated, while there was no significant change in the cell subsets of c-Met CAR-T(3G). The LDH release experiment showed that the kill efficiency of c-Met CAR-T(2G/3G) on SKOV-3 increased with the increase of effect-target ratio. When the effect-target ratio was 20:1, the kill efficiency of c-Met CAR-T(2G) reached (42.02 ± 5.17)% (P < 0.05), and the kill efficiency of c-Met CAR-T(3G) reached (51.40 ± 2.71)% (P < 0.05). ELISA results showed that c-Met CAR-T released more cytokine compared to CD19 CAR-T and activated T cells (P < 0.05). Moreover, the cytokine release of c-Met CAR-T(3G) was higher than c-Met CAR-T(2G) (P < 0.01). The CCK-8 results showed that after 48 h, the cell number of c-Met CAR-T(2G) was higher than that of c-Met CAR-T(3G) (P < 0.01). In conclusion, both the second and third generation c-Met CAR-T can target and kill c-Met-positive SKOV-3 cells, with no significant difference. c-Met CAR-T(2G) has stronger proliferative ability, and c-Met CAR-T(3G) releases more cytokines.
Humans ; Female ; Ovarian Neoplasms/immunology* ; Proto-Oncogene Proteins c-met/metabolism* ; Receptors, Chimeric Antigen/immunology* ; Cell Line, Tumor ; Cystadenocarcinoma, Serous/immunology* ; T-Lymphocytes/immunology*

OBJECTIVES: To explore the association of the expression of WW domain-containing ubiquitin E3 ligase 1 (WWP1) with immune infiltration in tumor microenvironment (TME) of ovarian cancer. METHODS: Ovarian cancer patient data from The Cancer Genome Atlas (TCGA) were used to analyze the association of WWP1 expression with patient prognosis. TISCH2 was utilized to analyze the changes in immune cell subtypes in TME of metastatic tumor and after chemotherapy. The impact of WWP1 on immune cell infiltration, somatic copy number alterations of WWP1 and evolution of immune cell subtypes was evaluated using TIMER and TIGER pseudo-time analysis. A deep learning model was used to analyze TCGA pathological images to investigate the effect of WWP1 on TME of ovarian cancer. RNA-seq analysis was conducted to identify the differentially expressed genes in WWP1-overexpressing SKOV3 cells and validate immune infiltration. Multicolor immunofluorescence assay was used to analyze the immune markers in SKOV3 and SKOV3/DDP cell xenografts in nude mice. RESULTS: The patients with high WWP1 expression levels had significantly lower overall survival rate (P=0.0012). High WWP1 expression levels and Stage IV disease were both associated with a poor prognosis (P<0.05). In metastatic ovarian cancer or after chemotherapy, the percentages of malignant tumor cells and tumor-associated fibroblasts increased in the TME, accompanied by elevated WWP1 levels. WWP1 expression level was positively correlated with pro-tumorigenic immunosuppressive cells (r=0.1323-0.3955, P<0.05) and negatively with tumor-inhibiting immune cells (r=-0.1949- -0.1333, P<0.05). Specific copy number alterations of WWP1 also influenced CD8⁺ T cell percentage and neutrophil infiltration levels in the TME. RNA-seq analysis of WWP1-overexpressing SKOV3 cells and immunofluorescence assay of the tumor-bearing mice yielded findings consistent with those of bioinformatics analysis. CONCLUSIONS WWP1 may serve as a prognostic biomarker and a potential target for immune regulation in the TME of ovarian cancer.
Female ; Ovarian Neoplasms/genetics* ; Humans ; Ubiquitin-Protein Ligases/metabolism* ; Tumor Microenvironment/immunology* ; Animals ; Mice ; Cell Line, Tumor ; Mice, Nude ; Prognosis ; Gene Expression Regulation, Neoplastic

Patients with ovarian cancer (OC) are often diagnosed at an advanced stage and have a poor prognosis because of extensive tumour metastasis. Tumour metastasis usually occurs in stages, which means that before the invasion of tumour cells, a pre-metastatic niche (PMN) has been formed to support the subsequent colonisation and growth of tumour cells. Tumour-associated macrophages (TAMs) are highly heterogeneous in terms of origin, phenotype and function. As one of the most abundant components in the tumour microenvironment (TME), TAMs can contribute to PMN formation by promoting organotropism, mediating immunosuppression, metabolic reprogramming, facilitating angiogenesis, and inducing cancer cell stemness. Therefore, TAMs play an important role in OC metastasis.
Humans ; Ovarian Neoplasms/immunology* ; Female ; Tumor-Associated Macrophages/immunology* ; Tumor Microenvironment ; Neoplasm Metastasis ; Animals ; Macrophages/metabolism*

Cold agglutinins(CA),autoantibodies against the antigen I or i on the surface of red blood cells,are mainly of IgM class,and the majority have κ light chains.They can lead to red blood cell agglutination at decreased body temperature and are usually associated with infections,drug reactions,autoimmune diseases,and hematological malignancies.However,solid tumors with CA are rare.We reported two cases of CA in the peripheral blood of patients with solid tumors.Peripheral complete blood cell count of the patients at admission showed reduced erythrocyte count and hematocrit,mismatching between erythrocyte count and hemoglobin,abnormally elevated levels of mean corpuscular hemoglobin and mean cell hemoglobin concentration.Peripheral blood smear showed erythrocyte aggregation.After the sample was preheated at 37 ℃ for 30 min,the reversibility of red blood cell aggregation was observed,and the erythrocyte parameters were corrected.
Humans ; Autoantibodies/isolation & purification* ; Female ; Breast Neoplasms/immunology* ; Ovarian Neoplasms/immunology*

Adult ; Female ; Humans ; Limbic Encephalitis ; etiology ; Ovarian Neoplasms ; complications ; Paraneoplastic Syndromes, Nervous System ; etiology ; Receptors, N-Methyl-D-Aspartate ; immunology ; Seizures ; etiology ; Teratoma ; complications

Acupuncture Therapy ; Edema ; etiology ; immunology ; therapy ; Female ; Humans ; Lower Extremity ; pathology ; Middle Aged ; Ovarian Neoplasms ; surgery ; Postoperative Complications ; etiology ; therapy

Primary retroperitoneal mucinous cystadenoma is an extremely uncommon tumor, even though mucinous cystadenoma often develops in the ovary and less frequently in the pancreas. A 21-year-old female was admitted to our hospital due to severe abdominal pain. A well-demarcated, oval shaped cystic tumor at the retropancreatic area with displacement of the pancreas and surrounding major vessels was observed on CT and MRI. Exploratory laparotomy was performed, and complete excision of the entire cyst was performed without complication. The pathologic finding was consistent with primary retropancreatic mucinous cystadenoma. To the best of our knowledge, this report is the first to describe a case of retropancreatic mucinous cystadenoma arising from the retropancreatic area in Korea.
Antibodies/metabolism ; Cystadenoma, Mucinous/*diagnosis/pathology/surgery ; Female ; Humans ; Magnetic Resonance Imaging ; Mucin 5AC/immunology ; Mucin-2/immunology ; Ovarian Neoplasms/*diagnosis/pathology/surgery ; Retroperitoneal Neoplasms/*diagnosis/pathology/surgery ; Tomography, X-Ray Computed ; Young Adult

OBJECTIVETo analyze the drift of T cell receptor (TCR) Vα and Vβ gene family CDR3 spectratype in response to ovarian carcinoma cells mediated by an anti-human ovarian carcinoma/CD3 bispecific single-chain antibody (BHL-1), and explore the mechanism of the bispecific single-chain antibody-mediated T cell immune response.

METHODSImmunoscopic spectratyping technique was used to analyze the TCR repertoire diversity (CDR3 spectratype distribution) of the T cells from 6 healthy donors before and after stimulation of the cells with human ovarian carcinoma in the presence of BHL-1. The predominant usage of TCR α and Vβ chain CDR3 was analyzed after the stimulation, and sequence analysis was performed for the CDR3 region of the monoclonal T cells.

RESULTSThe spectratypes of Vα and Vβ gene family TCR CDR3 region showed a Gaussian distribution before stimulation of the T cells from the 6 donors. After stimulation of the T cells, CDR3 spectratype drift occurred in the T cells, and some TCR Vα and Vβ families showed an anomalous and oligoclonal expansion. Different CDR3 sequences of the Vα and Vβ gene family TCR were found in the monoclonal T cells stimulated with BHL-1.

CONCLUSIONCDR3 spectratype drift occurs in TCR α and Vβ chains of T cells after stimulation with human ovarian carcinoma cells and BHL-1, indicating that the predominant usage of TCR Vα and Vβ families is associated with the specific T cell immune response mediated by BHL-1.

Antibodies, Bispecific ; immunology ; Cell Line ; Cell Line, Tumor ; Complementarity Determining Regions ; immunology ; Female ; Humans ; Monocytes ; immunology ; Ovarian Neoplasms ; immunology ; Receptors, Antigen, T-Cell, alpha-beta ; immunology ; Single-Chain Antibodies ; immunology

OBJECTIVETo examine the in vivo anti-metastatic effect of enhanced expression of CD40L cDNA in murine ovarian cancer OVHM cells (CD40L-OVHM) injected into the spleen on liver metastasis in mice.

METHODSOVHM cells were inoculated into the spleen of 6 to 8 week-old female B6C3F1 (C57BL/6N x C3H/He) mice. The established liver metastasis was identified by histopathology (HE staining). OVHM cells, DNA-pMKITneo-OVHM cells or CD40L-OVHM cells were inoculated into the spleen of female B6C3F1 mice and the expressions of CD11c in splenic cells were detected by flow cytometry. The specific cytotoxicity of splenic cells was detected by MTT assay, and the serum cytokines of IFN-gamma, TNF-alpha, IL-12, IL-4 and IL-10 of the mice were measured by enzyme linked immunoabsorbent assay. The liver metastases and the survival time of the mice were also recorded.

RESULTSThe mouse models with liver metastasis by injecting tumor cells into the spleen of mice were established. The expression of CD11c and the specific killing rate in CD40L-OVHM cells group was significantly higher than that in the OVHM cells group and DNA-pMKITneo-OVHM cells group. The expressions of IFN-gamma, TNF-alpha and IL-12 in the CD40L-OVHM cells group were much more increased than OVHM cells group and DNA-pMKITneo-OVHM cells group, but the expressions of IL-4 and IL-10 in the CD40L-OVHM cells group were decreased significantly (p < 0.05). The average weights of livers and spleens of mice in CD40L-OVHM cells group were significantly lower than those of DNA-pMKITneo-OVHM cells group and OVHM cells group. The survival time of mice in CD40L-OVHM cells group was also significantly longer than that in the OVHM cells group and DNA-pMKITneo-OVHM cells group (P < 0.05).

CONCLUSIONThe data directly demonstrate that the expression of CD40L in ovarian cancer cells (CD40L-OVHM) can enhance the proliferation and differentiation of dendritic cells in the spleen and induce specific cytotoxic effect of T cells in the spleen, and may regulate the immune function of peripheral blood cells and the immune balance between Th1 cells and Th2 cells, which maybe the possible mechanism induced by CD40L in mice inhibiting the development of liver metastasis.

Animals ; CD11c Antigen ; metabolism ; CD40 Ligand ; genetics ; metabolism ; Cell Differentiation ; Cell Line, Tumor ; Cell Proliferation ; DNA, Complementary ; genetics ; Dendritic Cells ; cytology ; Female ; Interferon-gamma ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-12 ; metabolism ; Interleukin-4 ; metabolism ; Liver Neoplasms ; pathology ; secondary ; Mice ; Mice, Inbred C3H ; Mice, Inbred C57BL ; Neoplasm Transplantation ; Ovarian Neoplasms ; metabolism ; pathology ; Spleen ; metabolism ; T-Lymphocytes, Cytotoxic ; immunology ; Tumor Necrosis Factor-alpha ; metabolism

Female ; Genes, p53 ; immunology ; Humans ; Ovarian Neoplasms ; genetics ; pathology ; Ovary ; pathology ; Proto-Oncogene Proteins ; immunology ; metabolism ; Proto-Oncogene Proteins p21(ras) ; Proto-Oncogenes ; immunology ; Sertoli-Leydig Cell Tumor ; genetics ; pathology ; ras Proteins ; immunology ; metabolism