RESUMEN
Recent studies have shown that cancer immunotherapy could be a promising therapeutic approach for the treatment of cancer. In the present study, to identify novel tumor-associated antigens (TAAs), the proteins expressed in a panel of cancer cells were serologically screened by immunoblot analysis and the eukaryotic elongation factor 2 (eEF2) was identified as an antigen that was recognized by IgG autoantibody in sera from a group of patients with head and neck squamous cell carcinoma (HNSCC) or colon cancer. Enzyme-linked immunosorbent assay showed that serum eEF2 IgG Ab levels were significantly higher in colorectal and gastric cancer patients compared to healthy individuals. Immunohistochemistry experiments showed that the eEF2 protein was overexpressed in the majority of lung, esophageal, pancreatic, breast and prostate cancers, HNSCC, glioblastoma multiforme and non-Hodgkin's lymphoma (NHL). Knockdown of eEF2 by short hairpin RNA (shRNA) significantly inhibited the growth in four eEF2-expressing cell lines, PC14 lung cancer, PCI6 pancreatic cancer, HT1080 fibrosarcoma and A172 glioblastoma cells, but not in eEF2-undetectable MCF7 cells. Furthermore, eEF2-derived 9-mer peptides, EF786 (eEF2 786-794 aa) and EF292 (eEF2 292-300 aa), elicited cytotoxic T lymphocyte (CTL) responses in peripheral blood mononuclear cells (PBMCs) from an HLA-A*24:02- and an HLA-A*02:01-positive healthy donor, respectively, in an HLA-A-restricted manner. These results indicated that the eEF2 gene is overexpressed in the majority of several types of cancers and plays an oncogenic role in cancer cell growth. Moreover, the eEF2 gene product is immunogenic and a promising target molecule of cancer immunotherapy for several types of cancers.
Asunto(s)
Antígenos de Neoplasias/genética , Quinasa del Factor 2 de Elongación/genética , Neoplasias/genética , Neoplasias/inmunología , Antígenos de Neoplasias/metabolismo , Línea Celular Tumoral , Quinasa del Factor 2 de Elongación/metabolismo , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Inmunoglobulina G/inmunología , Células MCF-7 , Neoplasias/patología , Análisis de Secuencia de ADN , Linfocitos T Citotóxicos/inmunologíaRESUMEN
Using a column-switching HPLC system combining a micro-ODS column and a chiral column, the amounts of D-proline (D-Pro) have been determined in 18 tissues, plasma and urine of mice. To avoid the enzymatic degradation of D-amino acids in vivo, a mutant mouse strain lacking D-amino acid oxidase activity (ddY/DAO(-) mouse) was used. In the brain, relatively large amounts of D-Pro were observed in the anterior pituitary, posterior pituitary and pineal glands. In the peripheral tissues, the amounts of D-Pro were high in the pancreas and kidney. Above all, it is surprising that the ddY/DAO(-) mice excreted large amounts of D-Pro in their urine (433 nmol/mL, 20 times that of L-Pro). The origin of D-Pro has also been investigated. By comparing germ-free mice and gnotobiotic mice, intestinal bacteria were shown to have no effect on the urinary D-Pro amount. Concerning the dietary origin, a notable amount of D-Pro was still excreted in the urine after starvation for 4 days, suggesting that some of the D-Pro is produced in the mice. Age-dependent changes in the urinary D-Pro amount have also been investigated from the postnatal 1st month up to 12 months, and ddY/DAO(-) mice were found to excrete large amounts of D-Pro in the urine constantly throughout their lives.
