CAR T cells redirected against tumor-specific antigen glycoforms: can low-sugar antigens guarantee a sweet success? / 医学前沿

Immune-based therapies have experienced a pronounced breakthrough in the past decades as they acquired multiple US Food and Drug Administration (FDA) approvals for various indications. To date, six chimeric antigen receptor T cell (CAR-T) therapies have been permitted for the treatment of certain patients with relapsed/refractory hematologic malignancies. However, several clinical trials of solid tumor CAR-T therapies were prematurely terminated, or they reported life-threatening treatment-related damages to healthy tissues. The simultaneous expression of target antigens by healthy organs and tumor cells is partly responsible for such toxicities. Alongside targeting tumor-specific antigens, targeting the aberrantly glycosylated glycoforms of tumor-associated antigens can also minimize the off-tumor effects of CAR-T therapies. Tn, T, and sialyl-Tn antigens have been reported to be involved in tumor progression and metastasis, and their expression results from the dysregulation of a series of glycosyltransferases and the endoplasmic reticulum protein chaperone, Cosmc. Moreover, these glycoforms have been associated with various types of cancers, including prostate, breast, colon, gastric, and lung cancers. Here, we discuss how underglycosylated antigens emerge and then detail the latest advances in the development of CAR-T-based immunotherapies that target some of such antigens.

Current status of bispecific antibody-mediated immunotherapy for hematologic malignancies / 中国输血杂志

Significant advances have been made in cancer immunotherapy recently, of which, bispecific antibodies (BsAbs), through bridging, redirecting and activating immune effector cells to kill cancer cells, are attracting increasing attention.Since the anti-CD19 and anti-CD3 BsAb, blinatumomab, was approved in 2014 by the FDA for the treatment of acute lymphoblastic leukemia, preclinical and clinical research with immune-cell-redirecting BsAbs have been fast growing in the area of hematologic malignancies. This review summarizes the current scientific and clinical investigation of BsAbs targeting different tumor-associated antigens from B lymphocytes, plasma cells and myeloid cells, covering three most common blood cancers, namely, lymphoma, multiple myeloma and leukemia. Further development for better therapeutic benefits and lower adverse events, are continuously being pursued, in particular, looking for more specific tumor antigens, optimizing antigen-antibody affinities, extending the half-life of BsAbs and redirecting different immune effector cells, whose breakthroughs and opportunities are soon to be delivered for the management of hematologic malignancies.

Serial Serum HER2 Measurements for the Detection of Breast Cancer Recurrence in HER2-Positive Patients / 한국유방암학회지

PURPOSE: The measurement of serum human epidermal growth factor receptor 2 (HER2) extracellular domain levels is a well-established method for evaluating whether a metastatic HER2-positive breast cancer patient will respond to HER2-targeted treatment. However, little is known about the value of serum HER2 for detecting disease relapse following curative surgical treatment in breast cancer patients. The purpose of this study was to evaluate the sensitivity of serum HER2, carcinoembryonic antigen (CEA), and carcinoma antigen 15-3 (CA 15-3) for the detection of disease recurrence in postoperative breast cancer patients with a primary HER2-positive tumor. METHODS: Serial measurements were taken of serum HER2, CEA, and CA 15-3 levels in patients with primary invasive HER2-positive breast cancer who underwent curative surgical treatment between January 2008 and December 2010. Following treatment, serum HER2 levels were monitored every 6 months using a chemiluminescence immunoassay. RESULTS: Overall, 264 patients were analyzed in this retrospective study. The median follow-up period was 27.7 months, and 24 patients relapsed during follow-up. The sensitivity of serum HER2, CEA, and CA 15-3 for the detection of disease recurrence was 37.5%, 25.1%, and 12.5%, respectively. Sensitivity increased to 45.8% when all three tumor markers were combined in the analysis. In a subgroup of patients without liver disease, the sensitivity of serum HER2, CEA, and CA 15-3 was 57.1%, 21.4%, and 14.3%, respectively. Of the 264 patients in this study, 80 patients had chronic hepatitis, liver cirrhosis, or abnormal aspartate aminotransferase or alanine aminotransferase levels during the follow-up period. Following the exclusion of these patients, the sensitivity of serum HER2 for the detection of disease recurrence increased to 57.1%. CONCLUSION: Serial serum HER2 measurement may be useful for the detection of disease relapse in patients with HER2-positive breast cancer. Abnormal liver function can result in elevated serum HER2 in the absence of disease recurrence.

Transfer of interleukin 6 gene into MCF-7 human breast cancer cells enhances expression of tumor-associated antigens / 中国临床药理学与治疗学

AIM: To investigate the mechanism by which IL-6 is involved in cancer prognosis, and further to demonstrate the relationship between IL-6 and tumor-associated antigens such as CA15-3, CEA and CA125 in breast cancer. METHODS: In the present study, we transfected an exogenous IL-6 gene into the MCF-7 cells. Secretion of CA15-3, CEA and CA125 into the culture media were measured by Enzyme Linked Immunosorbent Assay (ELISA). RESULTS: After a 72 hours in culture, the amount of IL-6 in the media of pCI-neo-IL-6-transfected MCF-7 cells (338.5±22.6 pg/106 cells) was significantly higher than that of non-transfected MCF-7 cells (25.4±4.6 pg/106 cells, P＜0.01, paired t-test), or pCI-neo-transfected MCF-7 cells (19.6±3.0 pg/106 cells, P＜0.01, paired t-test). The levels of CA15-3, CEA and CA125 secreted by the pCI-neo-IL-6-transfected MCF-7 cells were significantly higher than that of the parental MCF-7 cells or pCI-neo-transfected MCF-7 cells. The specific IL-6 antibody could decrease the expression of CA15-3, CEA and CA125 in both the MCF-7 cells and the IL-6 cDNA-transfected MCF-7 cells. CONCLUSION: Transfer of IL-6 gene augments tumor-associated antigens of human breast cancer cells. The association of elevated IL-6 concentration with a poor prognosis in cancer patients may partially be the result of increased expression of tumor-associated antigens by IL-6.

Advances in research of MAGE-A3 / 中国癌症杂志

Melanoma-associated antigen MAGE-A3 is expressed in various types of tumors,but not in normal cells except testis and placenta.MAGE-A3 can be presented by both HLA class I and HLA class II molecules after being processed into antigen peptide.MAGE-A3 has recently been widely used in the diagnosis and immunotherapy of tumors.This article reviews the applications of MAGE-A3 in the diagnosis and immunotherapy of tumors.

Active Immunization Study of Colon Cancer Derived 1-8D Peptide in HHD Mice

BACKGROUND: 1-8D gene is a member of human 1-8 interferon inducible gene family and was shown to be overexpressed in fresh colon cancer tissues. Three peptides 1-6, 3-5 and 3-7 derived from human 1-8D gene were shown to have immunogenicity against colon cancer. METHODS: To study tumor immunotherapy of three peptides we established an active immunization model using HHD mice. D(b-/-) x beta2 microglobulin (beta2 m) null mice transgenic for a chimeric HLA-A2.1/D(b-)beta2 m single chain (HHD mice) were challenged with B16/HHD/1-8D tumor cells and were immunized with irradiated peptide-loaded RMA- S/HHD/B7.1 transfectants. In therapy model tumor growth was retarded in HHD mice that were injected with 3-5 peptide-loaded RMA-S/HHD/B7.1. In survival test vaccination with 1-8D-derived peptide protects HHD mice from tumor progression after tumor challenge. RESULTS: These studies show that peptide 3-5 derived from 1-8D gene can be the most effective candidate for the vaccine of immunotherapy against colon cancer and highlight 1-8D gene as putative colon carcinoma associated antigens. CONCLUSION: We demonstrated that RMA-S/HHD/ B7.1 loaded with 1-8D peptides, especially 3-5, immunization generates potent antitumor immunity against tumor cells in HHD mice and designed active immunization as proper immunotherapeutic protocols.

The Clinical Utility of BTA TRAK, BTA stat, NMP22 and Urine Cytology in the Diagnosis of Bladder Cancer: A Comparative Study / 대한비뇨기과학회지

PURPOSE: Various tumor markers, developed recently for the diagnosis of bladder cancer, are reported to be superior to urine cytology in terms of their sensitivity and non-invasiveness. A prospective study was performed to evaluate the efficacies of BTA TRAK, BTA stat, NMP22 tests and urine cytology. MATERIALS AND METHODS: Between August 2001 and December 2001, 154 patients were involved in the study. Voided urine was used to perform BTA TRAK, BTA stat, NMP22 tests and cytology. The final diagnoses were pathologically proven transitional cell carcinoma (TCCa) of the bladder in 34 patients, gross or microscopic hematuria without detectable tumor in 27, history of TCCa without evidence of recurrence in 68 and benign urologic diseases in 25. RESULTS: The sensitivities of the BTA TRAK, BTA stat, NMP22 and urine cytology were 82.4, 79.4, 61.8 and 32.4%, respectively. The specificities were 75.8, 70.8, 95.8 and 92.5%, respectively. When the sensitivity was subdivided according to the tumor stage, grade and size, the bigger size yielded higher sensitivities in the NMP22 and BTA TRAK (p<0.01 and p=0.03, respectively). When the results of the urine cytology were combined with each of the other tests, the specificity was lower than each test alone for the BTA TRAK and BTA stat, while sensitivity and specificity were both higher than each test alone for the NMP22. CONCLUSIONS: The BTA TRAK, BTA stat and NMP22 tests are useful in overcoming the limitations of cystoscopy and cytology as the initial evaluation tools for patients where bladder cancer is suspected. In particular, the NMP22 may be more useful due to its enhanced specificity when combined with the urine cytology.

Establishment and biological characteristics of six human renal cell carcinoma cell lines / 中华泌尿外科杂志

Objective To establish human renal cell carcinoma (RCC) cell lines,and to investigate the cell phenotypes and expression of tumor associated antigens. Methods Fresh tumor tissues from pathologically proven human RCCs were primary cultured and passed generation to generation until a stably grow in vitro.The cell cloning form,chromosome and graft into nude mice in vivo were examined for cell lines.Immunofluorescent stain and flow cytometric analysis were carried out for cell's phenotype,RT PCR examination for MN/CA9 gene expression. Results Six human RCC cell lines have been established,including four of the cell lines derived from clear cells,one from mixed clear granular cells and one from papillary cells.All the cell lines showed the characteristics of malignant cells.All the lines highly expressed HLA ClassⅠ and HER2/neu.Three lines weakly expressed HLA ClassⅡ and one cell line highly expressed CD86 but all the lines did not express CD80.RT PCR showed that three cell lines have the expression of MN/CA9 gene. Conclusions These newly established RCC cell lines would provide a useful in vitro model for studies related to biological characteristics,tumor associated antigen,immunogenity and immunotherapy of human RCC.

The differential expression of tumor associated antigen CA-X in vitro and in vivo / 第二军医大学学报

Objective: To explore the differential expression of tumor associated antigen CA X in tumor cells in vitro and in vivo . Methods: Tumor xenografts of the human lung adenocarcinoma cell line SpcA1 were grown in athymia mice. The antigen was measured before and after xenograft by Western blotting and flow cytometric analysis. Results: The antigen CA X was a glycoprotein with a molecular weight of 170 000 to 200 000 . Human lung adenocarcinoma cell lines regain expression of antigen CA X when xenografted into athymia mice. Conclusion: These data indicate that CA X synthesis in human lung adenocarcinoma cells SpcA1 is regulated by factors presented only in the xenograft.

Tumor-associated antigen Ca - 125 in ovarian cancer.

Determinations of the serum Ca-125 levels were carried out on 45 samples from healthy women, 22 samples of benign ovarian tumors including tumor-like conditions, 33 samples of ovarian cancer and 14 cases of common epithelial cancer at second-look laparotomy. The Ca-125 levels were measured by enzyme immunoassay using a commercial kit by Fujirebio Inc. Japan. The cut off levels of Ca-125 values were set at > 30 IU/ml, which made the specificity, sensitivity and positive predictive values 80.0, 84.8 and 75.7 percent respectively. The demonstrable ovarian cancer gave positive Ca-125 values among serous cystadenocarcinoma in all case while endometrioid adenocarcinoma and mucinous cystadenocarcinoma were 80.0 and 66.6 percent respectively. Two sex cord tumors and 2 of 3 malignant germ cell tumors had positive Ca-125 values in low levels. Those with small residual cancer at second-look laparotomy, 1-2 cm in diameter, had positive Ca-125 values, and one of 12 patients had false-negative value despite the presence of microresidual cancer. Ovarian endometriosis had positive Ca-125 values about 55.5 percent but in low levels.

Purification of r-Glutamyltranspeptidase from Rat Primary Hepatoma Tissue and Preparation of a Tumor Associated Antigen

r-Glutamyltranspeptidase (r-GT) from a rat hepatoma induced by 3'-methyl-4-dimethylaminoazobenzene (3'-Me DAB) was purified 833 fold. The purified enzyme had a specific activity of 15.0 U/mg protein with an overall yield of 3.8%. The molecular weight of native r-GT was estimated as about 350,000 daltons, whichs a multicomplex of a single polypetide having a M W of 59,000. Anti r-GT rabbit antiserum cross-reacted with kidney r-GT as well as liver r-GT. Tryptic digestion of r-GT followed by separation with Con A sepharose column chromatography resulted in two major glycopeptides. A tumor associated antigen was prepared by the conjugation of a tryptic glycopeptide of r-GT to keyhole limpets hemocyanin and an antibody against this antigen cross-reacted preferentially with r-GT in rat hepatoma tissue.

Expression,purification and preparation of monoclonal antibody of HCA518 protein / 中国免疫学杂志

Objective; To express and purify HCA518 protein and prepare its monoclonal antibody ( McAb). Methods: The HCA518 protein was expressed with gene recombinant technique in prokaryotic system and purified with nickel chelate nitrilotriacetic acid(Ni-NTA) affinity chromatography column. Hybridoma cell lines that secreted anti-HCA518 McAb were established by cells fusion and screened by enzyme linked immunosorbent assay( ELISA). The specificity of anti-HCA518 McAb wa3 identified by Western blot assay. The HCA518 protein in tumor cells was stained by immunoflourescence assay. Results: Rcombinant HCA518 protein was expressed with a purity of 98%. Two hybridoma cell lines was selected and anti-HCA518 McAb was purified from mice ascites. The titers of anti HCA518 McAb in ascites were 1?10-4 and 5?10-4 respectively. The antibody belonged to IgG2b subtype and IgM. Anti-HCA518 McAb specifically reacted with recombinant HCA518 protein and tumor cells'nuclear protein (P100). The HCA518 protein was mainly located in cell nucleus. Conclusion: Stable hybridoma cell lines that secreted anti-HCA518 McAb have been established and anti HCA518 McAb was prepared with high specificity. It has important significance for detecting HCA518 protein in tumor tissues and determining malignant proliferation status of tumor cells and predicting its prognosis.