Screening of specific binding peptide targeting blood vessel of human esophageal cancer in vivo in mice / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Cancer of the esophagus and gastroesophageal junction remains a virulent malignancy with poor prognosis. Rapid progresses were made in chemotherapeutic agents and the development of molecular markers allowed better identification of candidates for targeted therapy. This study aimed to identify the candidate peptides used for anti-angiogenic therapy of esophageal cancer by in vivo screening C7C peptide library for peptides binding specifically to blood vessels of human esophageal cancer.</p><p><b>METHODS</b>The phage displayed C7C peptide library was injected intravenously into mice bearing human esophageal tumor xenografts under renal capsule. After 5 rounds of screening, 13 clones were picked up individually and sequenced. During each round of screening, titers of phage recovery were calculated from tumor xenograft and control tissues. Homing of these 9 peptides to tumor vessel was detected by calculating phage titers in the tumor xenograft and control tissues (lung and spleen) after each phage was injected into mice model, and compared with the distribution of phage M13 and VIII-related antigen in tumor xenograft by immunohistochemical staining. Comparisons among groups of data were made using one-way analysis of variance (ANOVA), followed by the Bonferroni multiple comparisons test.</p><p><b>RESULTS</b>The number of phage recovered from tumor tissue of each round increased gradually in tumor group while decreased in control groups (P < 0.01 in tumor and spleen, P < 0.05 in lung). Immunohistochemical staining showed similar staining pattern with M13 antibody or VIII-related antigen antibody, suggesting that phages displaying the selected peptides could home to blood vessel of human esophageal cancer. According to their DNA, 9 corresponding peptide sequences were deduced. And the homing ability to blood vessel of phages displaying the selected peptides was confirmed by comparing with their recovery in tumor and control tissues. Two motifs, YSXNXW and PXNXXN, were also obtained by analyzing the homology of these peptide sequences. The staining distribution of phage with the sequence of PNPNNST was similar to that of the blood vessel marker factor VIII-related antigen staining. After sequencing, each phage with the selected peptide of PNPNNST with 1.0 × 10(11) pfu/ml was injected intravenously into mice. The homing ability to tumor vessel of these 9 kinds of peptides in the xenograft was higher than control tissues (lung and spleen).</p><p><b>CONCLUSION</b>Nine peptides obtained from in vivo screening homed to the blood vessel of human esophageal cancer, and the two motifs of YSXNXW and PXNXXN are the possible biochemical recognition units binding to vascular endothelial cells of esophageal cancer.</p>

Detecting the activity of antibodies induced by recombinant TGFbeta1 vaccine / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To measure the neutralization activity in vitro of the antibodies induced by recombinant TGFbeta1 vaccine and to evaluate the vaccine's anti-liver fibrosis activity.</p><p><b>METHODS</b>Balb/c mice were immunized with a fusion protein of the human TGFbeta1 epitope-inserted into a hepatitis B core antigen using a prokaryotic expression system. The antibody produced by the recombinant vaccine was determined using ELISA. The biological activity of the anti-TGFbeta1 antibody induced by the vaccine was measured by MTT using mink lung epithelial cell Mv-1-Lu as inhibiting cells. The fusion protein was used as a vaccine in a mice hepatic-fibrosis model.</p><p><b>RESULTS</b>A high titer of anti-TGFbeta1 antibody and a low of anti-HBc antibody were detected in the mice after the immunization. The serum antibodies induced combined with the fusion and antigenic peptide prevented the TGFbeta1 inhibiting activity in the Mv-1-Lu cell.</p><p><b>CONCLUSION</b>Recombinant fusion protein can be used as a cytokine vaccine to induce high titers of anti-TGFbeta1 antibodies. Our results show the potentiality of the fusion protein to be used as a vaccine for preventing liver fibrosis.</p>

Fusion of TGF-beta 1 antigenic determinant and HBV core antigen as an anti-TGF-beta 1 vaccine / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To examine the expression and purification of the TGFbeta1 vaccine from prokaryotic expression system and to determine the antigenicity of the fusion protein of recombinant vector pET28a/ HBcAg1-71-TGFbeta132-HBcAg89-144.</p><p><b>METHODS</b>The reconstructed vector pGEMEX-1/CTC was subcloned to pET28a and transformed into E. coli BL21 (DE3). The recombinant 6xHis- HBcAg1-71- TGFbeta132- HBcAg89-144 was to be expressed after induction by IPTG and purified with Ni-NTA-His affinity chromatography. The detection of the formation of core-like particles was done under an electron microscope and of their antigenity by using ELISA and Western blot.</p><p><b>RESULTS</b>A 2.46 x 10(4) protein was obtained by optimizing the conditions for both expression and purification. The protein had the TGFbeta1 antigenicity but not a HBc antigenity and the formed core-like particles were bigger than natural core particles.</p><p><b>CONCLUSION</b>The recombinant fusion protein in the prokaryotic expressed system can be used as an anti-TGFbeta1 vaccine to inhibit hepatic fibrosis.</p>

Osteopontin expression and its relation to invasion and metastases in gastric cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the correlation between expression of the osteopontin (OPN) and invasion and metastases in gastric cancer.</p><p><b>METHODS</b>The expression of OPN, NF-kappaB p65 and matrix metallo-proteinase 9 (MMP-9) was detected by immunohistochemistry in non-cancer gastric tissue (n = 12 cases) and gastric cancer tissue (n = 72 cases).</p><p><b>RESULTS</b>(1) OPN, NF-kappaB p65 and MMP-9 were not expressed in 12 non-cancer gastric tissue samples(group A). Their expression rates were 43.3%, 40.0% and 46.7% respectively in 30 gastric cancer samples without lymph nodes metastasis (group B), but they increased to 76.9%, 73.1% and 80.8% in 26 gastric cancer samples with lymph nodes metastases (group C), and 87.5%, 81.3% and 93.8% respectively in 16 gastric cancer samples with lymph node and distant metastases (group D). (2) There were statistically significant differences in their expressions between group D and group B (P(a) = 0.004, P(c) = 0.007, P(e) = 0.002), and between group C and group B (P(b) = 0.011, P(d) = 0.013, P(f) = 0.009). (3) Despite some differences in positive expression rates, correlations existed between OPN and NF-kappaB p65, and between NF-kappaB p65 and MMP-9 (P(1) = 0.042, P(2) = 0.013; r(1)= 0.67, r(2)= 0.72).</p><p><b>CONCLUSION</b>Osteopondin espression is closely related to the invasion and metastases of gastric cancer. It may upregulate the expression of metastasis-related molecule MMP-9 by activating NF-kappaB pathway.</p>