OBJECTIVESTo obtain a single-chain antibody with high affinity against hepatocellular carcinoma (HCC).

METHODSA second single-chain antibody mutant library was established using an error-prone PCR and a phage display. Single-chain antibodies with high affinity for hepatocellular carcinoma were selected using ELISA.

RESULTSThe content of the second single-chain antibody mutant library was about 4.5 x 10(7). Two selected mutants, M90 and M116, were obtained after 3 rounds of panning and ELISA. Immunoassay showed that both M90 and M116 could bind to human HCC cells. The relative affinity of M90 was 1.7-fold higher than that of the original antibody, and M116 was 2-fold higher than that of the original antibody.

CONCLUSIONError-prone PCR is an effective and simple method for affinity maturation of antibodies isolated from a phage antibody library.

Antibodies, Neoplasm ; immunology ; Antibody Affinity ; Antibody Specificity ; Carcinoma, Hepatocellular ; immunology ; pathology ; Humans ; Immunoglobulin Fragments ; immunology ; Immunoglobulin Variable Region ; genetics ; immunology ; Liver Neoplasms ; immunology ; pathology ; Mutation ; Peptide Library

BACKGROUND: It is well known that both gastric and intestinal phenotypic cell markers are expressed in gastric cancers. This study was aimed at investigating the correlation between gastric and intestinal phenotypic marker expression patterns of tumors and the clinicopathologic characteristics of gastric carcinomas. METHODS: We evaluated phenotypic marker expression by immunohistochemical staining with monoclonal antibodies. All tumors were classified as gastric (G), gastric and intestinal mixed (GI), intestinal (I), or null (N) phenotype. RESULTS: The tumors were phenotypically divided into G-phenotype tumors (33.2%), GI-phenotype tumors (25.7%), I-phenotype tumors (26.8%), and N-phenotype tumors (14.3%). N-phenotype tumors were associated with more corporeal location than GI- and I-phenotype tumors (p=0.009 and p=0.007, respectively), a larger size than I-phenotype tumors (p=0.007), a higher proportion of advanced gastric cancers than G-, GI-, and I-phenotype tumors (p=0.003, p< 0.001, and p< 0.001, respectively), more perineural invasion than G-, GI-, and I-phenotype tumors (p=0.076, p=0.003, and p=0.003, respectively), and more lymph node metastasis than GI-phenotype tumors (p=0.017). I-phenotype tumors were associated with a higher proportion of intestinal-type tumors than G-, GI-, and N-phenotype tumors (p< 0.001, p=0.011, and p< 0.001, respectively). CONCLUSION: Our results indicate that the gastric and intestinal phenotypic marker expression pattern of tumors is prognostically useful for patients with gastric carcinoma.
Tumor Markers, Biological/genetics ; Stomach Neoplasms/genetics/immunology/*pathology ; Prognosis ; *Phenotype ; Middle Aged ; Male ; Intestinal Neoplasms/genetics/immunology/*pathology ; Immunohistochemistry ; Humans ; Female ; Carcinoma/genetics/immunology/*pathology ; Antibodies, Neoplasm ; Antibodies, Monoclonal

OBJECTIVETo construct a adevoviral vector harboring human renal tumor-associated antigen G250 gene for transfecting the dendritic cells (DCs) and treating renal tumors.

METHODSThe G250 genes were cloned into the shuttle plasmid pDC316 to construct pDC316-G250, which was cotransfected with the rescue plasmid pBHGlox(delta)E1,3Cre into 293 cells to obtain the recombinant adenovirus Ad/G250. The inserted gene and its expression were identified by RT-PCR and fluorescence-activated cell sorting (FACS) after recombinant adenovirus transfection of the DCs. The recombinant adenoviral vector was purified by CsCl banding and titrated by TCID50.

RESULTS AND CONCLUSIONThe recombinant adenoviral vector of G250 gene was successfully constructed and high titer of the recombinant adenoviruse was obtained. G250 mRNA and protein expressions were identified in Ad/G250-transfected DCs. The titer of the virus stocks reached 5.6x10(9) IU/ml.

Adenoviridae ; genetics ; Antigens, Neoplasm ; biosynthesis ; genetics ; Carcinoma, Renal Cell ; genetics ; immunology ; pathology ; Dendritic Cells ; metabolism ; Genetic Vectors ; genetics ; Humans ; Kidney Neoplasms ; genetics ; immunology ; pathology ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection ; Tumor Cells, Cultured

Cancer cell vaccine-based immunotherapy has received increasing interest in many clinical trials involving patients with breast cancer. Combining with appropriate adjuvants can enhance the weak immunogenic properties of tumor cell lysates (TCL). In this study, diphtheria toxin (DT) and two tandem repeats of mycobacterial heat shock protein 70 (mHSP70) fragment 407-426 (M2) were conjugated to TCL with glutaraldehyde, and the constructed cancer cell vaccine was named DT-TCL-M2. Subcutaneous injection of DT-TCL-M2 in mice effectively elicited tumor-specific polyclonal immune responses, including humoral and cellular immune responses. High levels of antibodies against TCL were detected in the serum of immunized mice with ELISA and verified with Western blot analyses. The splenocytes from immunized mice showed potent cytotoxicity on Ehrlich ascites carcinoma cells. Moreover, the protective antitumor immunity induced by DT-TCL-M2 inhibited tumor growth in a mouse breast tumor model. DT-TCL-M2 also attenuated tumor-induced angiogenesis and slowed tumor growth in a mouse intradermal tumor model. These findings demonstrate that TCL conjugated with appropriate adjuvants induced effective antitumor immunity in vivo. Improvements in potency could further make cancer cell vaccines a useful and safe method for preventing cancer recurrence after resection.
Adjuvants, Immunologic ; Animals ; Bacterial Proteins ; genetics ; immunology ; Cancer Vaccines ; immunology ; Carcinoma, Ehrlich Tumor ; immunology ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Diphtheria Toxin ; genetics ; immunology ; Female ; HSP70 Heat-Shock Proteins ; genetics ; immunology ; Immunoglobulin G ; immunology ; Immunotherapy ; Mice ; Neovascularization, Pathologic ; Peptide Fragments ; genetics ; immunology ; Recombinant Fusion Proteins ; genetics ; immunology ; T-Lymphocytes, Cytotoxic ; immunology ; Tandem Repeat Sequences

OBJECTIVETo investigate apoptosis of tumor infiltrating dendritic cells (TIDC) and their expression of Fas/FasL (CD95/CD95L) in human endometrioid adenocarcinoma.

METHODSThe apoptotic rate of TIDC was measured in 45 cases of endometrioid adenocarcinoma and 20 cases of normal endometrium tissues (control) by double-label immunohistochemistry using the monoclonal antibody S-100 protein and TUNEL technique. The expressions of Fas and FasL in TIDCs were detected using double-label immunohistochemistry and imaging analysis.

RESULTSThe apoptotic rate of TIDCs in endometrioid adenocarcinoma were significantly higher than that in normal endormetrium [(13.02∓0.64)% vs (6.82∓0.53)%, P<0.05]. The expression levels of Fas in the TIDCs were significantly lower, whereas FasL expression significantly higher in endometrioid adenocarcinoma than in normal endormetrium (7.88∓1.05 vs 19.25∓3.03, P<0.05; 12.95∓2.25 vs 7.51∓1.14, P<0.05).

CONCLUSIONIncreased apoptosis of the TIDCs and abnormal expression of Fas/FasL in TIDCs in endometrioid adenocarcinoma may lead to tumor immune escape.

Apoptosis ; physiology ; Carcinoma, Endometrioid ; immunology ; metabolism ; pathology ; Case-Control Studies ; Dendritic Cells ; immunology ; Endometrial Neoplasms ; immunology ; metabolism ; pathology ; Fas Ligand Protein ; genetics ; metabolism ; Female ; Humans ; Lymphocytes, Tumor-Infiltrating ; immunology ; Tumor Escape ; fas Receptor ; genetics ; metabolism

OBJECTIVETo investigate the effect of anti-ICAm-1/LAF-1 McAb on the cytotoxicity of tumor infiltrating lymphocytes from oral squamous cell carcinoma.

METHODSTIL were isolated from fresh tumor tissues in 15 patients with oral squamous cell carcinoma and cultured in vitro. The expression of ICAM-1/LAF-1 in fresh and activated TIL was examined by FACS. The cytotoxicity of TIL against auto tumor cells and the level of TNF-alpha produced by TIL were compared before and after interrupted with anti-ICAM-1/LAF-1 McAb.

RESULTSThe expression of ICAM-1 and LAF-1 in the activated TIL significantly increased comparing with fresh TIL (P < 0.05). Interrupted by anti-ICAM-1 McAb, the cytotoxicity of TIL against auto tumor cell was not significantly changed. Interrupted by anti-ICAM-1 and anti-LAF-1 McAb, the level of TNF-alpha produced by TIL showed no differences comparing with the control (P < 0.05), but the cytotoxicity of TIL against auto tumor cells significantly decreased (P < 0.05).

CONCLUSIONThe effect of anti-LAF-1 McAb may result from steric interference of antigen-nonspecific adhesion process.

Antibodies, Monoclonal ; pharmacology ; Carcinoma, Squamous Cell ; immunology ; pathology ; Cells, Cultured ; Cytotoxicity, Immunologic ; Female ; Humans ; Intercellular Adhesion Molecule-1 ; biosynthesis ; genetics ; Lymphocyte Function-Associated Antigen-1 ; biosynthesis ; genetics ; Lymphocytes, Tumor-Infiltrating ; immunology ; pathology ; Male ; Mouth Neoplasms ; immunology ; pathology

Antibodies, Neoplasm ; blood ; immunology ; Carcinoma, Hepatocellular ; pathology ; Cell Differentiation ; Female ; Humans ; Liver Neoplasms ; pathology ; Male ; Tumor Suppressor Protein p53 ; blood ; genetics ; immunology

OBJECTIVETo study the clinicopathologic features and immunophenotype of renal cell carcinomas, and to discuss their diagnostic value.

METHODSThe clinicopathologic features of 114 cases of renal cell carcinoma were reviewed and categorized on the basis of 2004 WHO classification. Immunohistochemical study for a panel of antibodies (including CK, CD10, vimentin, CD117, AMACR, CK7 and TFE3) was carried out. The follow-up data, if available, were also analyzed.

RESULTSThe cases were reclassified into 5 subtypes, including 77 cases (67.5%) of clear cell carcinoma (CCRCC), 11 cases (9.6%) of papillary renal cell carcinoma (PRCC), 14 cases (12.3%) of chromophobe renal cell carcinoma (chrRCC), 10 cases (8.8%) of renal carcinoma associated with Xp11.2 translocations/TFE3 gene fusions (Xp11.2RCC) and 2 cases (1.8%) of unclassified renal cell carcinoma (unRCC). Immunohistochemical study showed that the expression rates of CK, CD10 and vimentin in CCRCC were 93.5% (72/77), 93.5% (72/77) and 75.3% (58/77), respectively. On the other hand, all the 11 cases of PRCC studied were positive for AMACR. The expression rate of CD117 in chrRCC was 78.5% (11/14). In the 10 cases of Xp11.2 RCC studied, the expression rates of TFE3, AMACR, CD10 and CK were 100% (10/10), 100% (10/10), 90% (9/10) and 70% (7/10), respectively.

CONCLUSIONSThe various subtypes of renal cell carcinomas are heterogeneous in histologic appearance and demonstrate distinctive immunophenotype. The expressions of CD10, vimentin, CD117, AMACR, CK7 and TFE3 are helpful in the differential diagnosis.

Adenocarcinoma, Clear Cell ; pathology ; Adolescent ; Adult ; Aged ; Aged, 80 and over ; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors ; genetics ; immunology ; metabolism ; Biomarkers, Tumor ; analysis ; genetics ; Carcinoma, Papillary ; immunology ; pathology ; Carcinoma, Renal Cell ; immunology ; metabolism ; pathology ; Female ; Gene Fusion ; Humans ; Immunophenotyping ; Kidney Neoplasms ; immunology ; metabolism ; pathology ; Male ; Middle Aged ; Neprilysin ; analysis ; Racemases and Epimerases ; analysis ; genetics ; Translocation, Genetic ; Vimentin ; analysis ; World Health Organization ; Young Adult

Carcinoembryonic Antigen ; metabolism ; Carcinoma in Situ ; immunology ; pathology ; virology ; Carcinoma, Ductal, Breast ; immunology ; pathology ; virology ; Cervical Intraepithelial Neoplasia ; immunology ; pathology ; virology ; DNA, Viral ; analysis ; Diagnosis, Differential ; Female ; Human papillomavirus 16 ; genetics ; isolation & purification ; Humans ; Ki-67 Antigen ; metabolism ; Uterine Cervical Dysplasia ; immunology ; pathology ; virology ; Uterine Cervical Neoplasms ; immunology ; pathology ; virology

Brain metastasis was a common metastasis site and leading cause of death in non-small cell lung cancer (NSCLC). Tyrosine kinase inhibitors had improved survival of NSCLC patients with positive drive gene. It also brings good news to NSCLC patients with positive drive gene and brain metastases. However, there is still no effective treatment for NSCLC patients with drive gene-negative and brain metastases. In recent years, immunotherapy has made breakthrough progress and become important first and second line treatment options of NSCLC especially in patients with drive gene-negative. The role of immunotherapy in specific populations of NSCLC-brain metastasis patients, especially drive gene-negative patients has become the focus of attention. In this report, we review the research progress of immunotherapy in NSCLC with brain metastases, especially in driver-negative patients, analyze the limitations of existing research and future challenge.
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Brain Neoplasms ; immunology ; secondary ; therapy ; Carcinoma, Non-Small-Cell Lung ; genetics ; pathology ; Humans ; Immunotherapy ; methods ; Lung Neoplasms ; genetics ; pathology ; Patient Selection