Although some evidence has been documented on EGFR/PI3K mediation of Akt activation in breast cancers, ILK and DNA-PK have not been investigated so far. The aim of this study was to analyze the expression of phosphorylated Akt (pAkt) in breast cancer, with respect to its upstream regulators. The immunostaining of pAkt (Ser473) in 70 invasive breast cancers revealed that status of CerbB2 could play a major role in Akt phosphorylation, while ILK was also involved in the stimulated level of pAkt. The results would provide an important clue for the activation of Akt and potential targeted therapy in breast cancer.

In a previous study, we demonstrated that telomerase transcriptional elements-interacting factor (TEIF) could up-regulate the expression of telomerase and DNA polymerase beta, increasing resistance to genotoxic agents. Here, we further report that TEIF can be stimulated by DNA damage and we have identified a cluster of repeated polypyrimidine tracts in the promoter of TEIF, which mediate both its basal transcription and its response to genotoxic agents. These polypyrimidine tracts are arranged in three types of repeating units and in each of these units there are 14 bp length tandem sequences, which are repeated three times. These sequences are also characteristically separated by an 11 bp interval sequence. Among these units, one type (5'-CCCCCCCATCCCCG-3') has been found to be involved in the transcriptional regulation of TEIF. At the same time, PTB1 (polypyrimidine tract-binding protein 1) has been shown to repress TEIF expression through interaction with this element. Up-regulation of TEIF may be achieved by PTB1 suppression that is induced by DNA damage, or by an olignucleotide decoy, which mediates reversal of suppression. This study provides new insight into the mechanism through which TEIF is involved in DNA damage response, together with insight into the role of polypyrimidine tracts in transcription regulation.

Objective:To get insight into the molecular features in tumorigenesis of bone tumors,the changes in telomeric length, expression of binding proteins and telomerase as well as tumor-associated gene p53 and c-myc were analyzed.Methods:The telomere length in 20 osteosarcomas,25 chondrosarcomas,14 fibrous dysplasia of bone were measured by telomere fluorescence in situ hybridiza- tion(Telo-FISH),and the expression ofTRF1,POT1,hTERT,p53 or c-myc was analyzed by immunohistochemistry.In situ hybridization was also used to detect expression of hTERT.Results:The average telomeri length in 20 cases of osteosarcomas were determined 0.31, 0.41 for 25 of ehondrosareomas,0.52 for 14 of fibrous dysplasia of bone.There was a significant difference in the three groups of tumors in statistics(P0.05).The expression of YRF1 or POT1 in osteosarcoma or chondrosarcoma was markedly weaker than that in fibrous dysplasia of bone(P0.05).In contrast,hTERT in osteosarcoma and chondrosarcoma was much stronger(P0.05).The nuclear staining of P53 and expression ofc-myc were far more frequent in osteosarcoma and chondrosarcoma(P0.05).In addition,the telomeric length in bone tumors was negatively with the expression of telomere binding protein TRF1 and POT1,but was positively correlated with ex- pression of hTERT,nuclear accumulation of P53 and c-myc staining.Conclusions:The telomitic length reflects the malignant phenotype and the shortening of telomere correlates with gene mutation of bone tumor.

OBJECTIVE To explore the significance in the change of telomere length in mesenchymal sarcomas, through analyzing telomere length and expression of its associated proteins, including TRF1, POT1, hTERT, P53 and c-myc. METHODS The telomere length in 20 cases of osteosarcomas, 25 of chondrosarcomas, 19 of rhabdomyosarcomas, 26 of liposarcomas was measured by telomere fluorescence in situ hybridization (Telo-FISH), and the expression of TRF1, POT1, hTERT, p53 or c-myc was analyzed by immunohistochemistry, respectively. RESULTS The telomere length in osteosarcomas was significantly shorter than that of either chondrosarcomas or liposarcomas (P<0.05). Similarly, the telomere length of rhabdomyosarcoma was shorter than that of chondrosarcoma (P<0.05). Meanwhile, telomere shortening was positively correlated with down expression of telomere binding proteins TRF1 and POT1 (P<0.05), but trends were detected more frequently in positive expression of hTERT (P<0.05) and in nuclear accumulation of P53 or expression of c-myc. With advancing in histological grading, telomere length was shortened markedly in chondrosarcomas, especially in liposarcomas (P<0.05). CONCLUSION The shortening of telomere could prevail in mesenchymal sarcoma and reflect the malignant potential. Telomere attrition usually correlated with down expression of POT1, TRF1 and with increased levels of hTERT, P53 and c-myc.

OBJECTIVE:To evaluate the expression of TEIF protein in human tumors of soft tissue and its significance. METHODS:Anti-TEIF polyclonal antibody was generated by immunization of E.coli expressed His-TEIF protein. The expression of TEIF in 166 cases of sarcomas and 28 case benign tumors or tumor-like lesions of soft tissue arranged in tissue chip was analyzed by immunohistochemistry. RESULTS:Polyclonal antibody obtained from immunized rabbit was verified in Western blot to prove its specific reactivity with native TEIF protein. The immunohistochemical staining of TEIF showed that about 58% (97/166) of sarcomas were positive and significantly different from that of benign tumors or tumor-like lesions (11%, 3/28). The positive staining was predominantly in synovial sarcoma 94% (16/17), primitive neuroectodermal tumor (PNET) 91% (21/23), both of which were significantly higher than 43% (6/14) of dermatofibrosarcoma protuberans, 38% (6/16) of myxofibrosarcoma, 36% (8/22) of malignant peripheral nerve sheath tumor, 32% (6/19) of liposarcoma, (P < 0.05, respectively), but not higher than 75% (15/20) of malignant fibrous histiocytoma, 70% (7/10) of rhabdomyosarcoma or 64% (9/14) of leiomyosarcoma. Meanwhile, strong positive staining of TEIF (>or= 2+) was frequently observed in PNET (83%, 19/23) and synovial sarcoma (76%, 13/17). With respect to FNCLCC grading, 19 cases of grade I sarcoma TEIF was 32% (6/19) and strong positive was 11% (2/19), 44 cases of grade II sarcoma was 48% (21/44) and 32% (14/44), and 70 of grade III was 84% (59/70) and 70% (49/70). The rate of either positive or strong positive in grade III sarcoma was significantly different from that of either grade I or II (P < 0.05), but no difference between the latter two groups (P > 0.05). CONCLUSIONS:TEIF protein could be detected in large part of soft tissue sarcomas, and it not only over-expressed in most of PNET, synovial sarcomas, but also correlated with histological grading.

Abstract： Centrosome amplification, a characteristic of cancer cells, has been believed a driving force for genetic instability in the development of cancer and received extensively exploration in recent years. In our previous work, we have demonstrated that TEIF（transcriptional elements-interacting factor）distributes at the centrosome and regulates its status in physiological condition, DNA damage response or carcinogenesis. Here, we further expand our knowledge of TEIF to the downstream effector of EGF/PI3K/Akt signaling. The addition of EGF or transfection of active Akt stimulated TEIF expression, resulting in an increase of centrosome splitting and amplification, while the inhibitors of either PI3K or Akt attenuated the changes in TEIF and its associated centrosome status. A consensus motif for Akt phosphorylation (RHRVLT) was revealed in TEIF domain of localizing at the centrosome and identified as the site for phosphorylation of TEIF both in vitro and in vivo experiments, indicating TEIF as a direct substrate to Akt1. Furthermore, in a collection of aggressive breast cancers a positive correlation between the expression of TEIF and centrosome amplification was clarified and also closely concerned to the over-expression of CerbB2 (a member of EGFR family) and the level of phosphorylated Akt (pAkt) in cancer tissues. These findings not only could link EGF/PI3K/Akt oncogenic signaling with centrosome amplification, but also could provide more choices in the development of specific inhibitors for anti-cancer therapeutic agents targeting to EGF/PI3K/Akt signaling.