| Background&Objective:Breast cancer, one of the most common primary malignant carcinomas in the world, has increased annually. Because breast cancer can remain undetected for a long period of time resting in a poor prognosis, there is an urgent need to identify and treat this cancer as early as possible. At present, breast cancer detection is addressed by different diagnostic techniques. The main treatment for breast cancer includes surgical management, radiotherapy, chemotherapy, endocrine therapy and immunotherapy.The major obstacles impeding current cancer therapies are side effects and low specificity. One way to improve the efficacy of therapeutic agents is to selectively target the tumor site, thereby sparing normal tissues. The development of targeted therapeutic methodologies relies, in most cases, on the availability of agents that specifically bind to tumor-associated markers. The display of polypeptides on the surface of filamentous phage, together with an efficient selective-amplification of the desired binding specificities using affinity capture, represents an efficient route towards the isolation of specific peptides and proteins that could act as vehicles for a tumor targeting application.In this study, we selected a small peptide from a12-mer peptide library displayed on the surfaces of filamentous phages in TA Ⅱ mice, which targets breast cancer cells. A single-phage clone was identified by immunostaining showing affinity for breast cancer tissues.Methods:In the first part, a mouse model for carrying breast cancer xenografts was established using Tientsin Albinao Ⅱ mice(TA11). A12-peptide library was biopanned through4rounds. Phages were recovered and titrated from tumor xenografts and control tissue (liver).In the second part, we selected ten clones and sequenced. A peptide sequence was obtained by company. At the same time, the distribution of phages was detected by immunohistochemical staining.Results:The frist part:1.1.0-cm-sized solid tumors were established in the TA Ⅱ mice over a period of4weeks. Pathological examinations of the sections with H&E stain showed hyperplastic cancer cells irregularly arrange in a tubular form, which is consistent with the characteristics of breast cancer.2. After4rounds of successive in vivo selection, phage homing to the breast cancer in the fourth round was4.1times than of the first cycle.3. Phage homing to the breast cancer xenografs was enriched,14-fold compared to that recovered from liver tissue, showing that by the fourth round binding of the phage to the breast cancer tissues was significantly increased.The second part1. After analyzing the sequence of the inserted peptide, results showed that ASANPFPTKALL appeared8times and PSRNPFPTKALL appeared2times. In conclusion, ASANPFPTKALL maybe the special peptide binding to breast cancer tissues.2. Immunohistochemical staining results indicated that the selected phage binding was found on cell membranes and in the cytoplasm of the tumor cells(murine or Homo sapien).3. Compared with control tissues,G1is the peptide which is specially bond to breast cancer cells.4. The peptide, ASANPFPTKALL, may have significant potential as a tumor-targeting and tumor-imaging agent.Conclusions:1. The phage clones displaying the peptide sequence ASANPFPTKALL were found bond to breast cancer cells after four rounds in vivo.2. Immunohistochemical staining results indicated that the selected phage binding was found on cell membranes and in the cytoplasm of the tumor cells(murine or Homo sapien).3. Compared with control tissues,G1is the peptide which is specially bond to breast cancer cells.4. Tumor-specific homing peptides may provide an effective tool for breast cancer target therapy. The in vivo phage display selection technique employed in this study was feasible and applicable to screening peptides that home to breast cells. |
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