Novel affibody molecules as potential agents in molecular imaging for MAGE-A3-positive tumor diagnosis.

BACKGROUND: The cancer-testis protein melanoma antigen A3 (MAGE-A3) is highly expressed in a broad range of malignant tumor forms. It has been confirmed that affibody molecules, a novel family of small (∼6.5 kDa) targeting proteins, are useful agents for molecular imaging and targeted tumor treatment. As a novel agent for in vivo molecular imaging detection of MAGE-A3-positive tumors, the efficacy of affibody molecules was assessed in this research. METHODS: In this study, three cycles of phage display library screening resulted in the isolation of two new affibody molecules (ZMAGE-A3:172 and ZMAGE-A3:770) that attach to MAGE-A3. These molecules were then expressed in bacteria and purified. The affibody molecules with high affinity and specificity were evaluated using western blotting, immunohistochemistry, indirect immunofluorescence, surface plasmon resonance, and near-infrared optical imaging of tumor-bearing nude mice. RESULTS: The selected ZMAGE-A3 affibodies can precisely bind to the MAGE-A3 protein in living cells and display high-affinity binding to the MAGE-A3 protein at the molecular level. Furthermore, the accumulation of DyLight755-labeled ZMAGE-A3:172 or ZMAGE-A3:770 in MAGE-A3-positive tumors was achieved as early as 30 min and disappeared at 48 h post-injection. CONCLUSION: Our findings support the potential of the two MAGE-A3 protein-binding affibody molecules for their use as molecular imaging agents.

Noncystoscopic removal of retained ureteral stents in children: A retrospective study from a single-center.

Cystoscopic technique is the current common method of retrieving double J ureteral stent in most pediatric urological centers. In this study, we evaluated the feasibility and efficacy of a novel noncystoscopic method to remove retained ureteral stents in pediatric patients.We reviewed all medical records from a total of 102 patients who were treated in our hospital between January 2013 and December 2016 to remove the double J ureteral stent retained into the ureter. The pediatric patients were divided into 2 groups based on different surgical options: cystoscopic group and noncystoscopic group. The surgery time (including time for instrument preparation), operation time, expenses, postoperative urination discomfort, and hospitalization were compared between the 2 groups.The noncystoscopic group took significantly less time for surgery and operation than the cystoscopic group (surgery time:7.40 ±â€Š3.75 vs 18.42 ±â€Š2.77 min, P <.05; operation time: 3.54 ±â€Š2.03 vs 4.48 ±â€Š2.04 min, P <.05). The mean spending for patients in the noncystoscopic group were less than that in the cystoscopic group ($736.70 ±â€Š105.96 vs $618.23 ±â€Š110.31, P <.05). There were less children with postoperative urination discomforts in the noncystoscopic group than that in the cystoscopic group (8 vs 20 cases, χ = 4.241, P <.05). The mean hospitalization of the noncystoscopic group was shorter than that of the cystoscopic group (3.20 ±â€Š1.25 vs 4.13 ±â€Š1.63 d, P <.05). The differences in all comparison projects were significant.The noncystoscopic procedure is a safe and viable technique that may be used successfully in pediatric urology. This novel procedure which is much safer and more affordable provides an alternative solution to remove retained ureteral stents in child patients.

Induction of Humoral and Cellular Immune Responses in Mice by Multiepitope Vaccines Composing of Both T and B Lymphocyte Epitopes of MAGE-A3 which are Recombined into HBcAg.

BACKGROUND: Melanoma-associated antigen-A3 (MAGE-A3) is a tumor specific antigen and a potential candidate for cancer immunotherapy. We had screened three immunodominant multiepitopes of MAGE-A3, and identified these multiepitope peptides had significantly higher reactivity to serum samples from gastric cancer patients. However, the immune responses of three multiepitope peptides carried by HBcAg in mice have not been investigated. OBJECTIVES: The main objective of this study was to analyze the humoral and cellular immune responses in mice induced by these three multiepitope vaccines of MAGE-A3. METHODS: Three multiepitopes of MAGE-A3 (MAGE-A3(EPI-1, or -2, or -3)) were respectively inserted at HBcAg major immunodominant region (HBcAg(MIR)) of the pET21a(+)/HBcAg(MIR) recombinant plasmid. These recombinant chimeras were identified by PCR, and transfected respectively into E. Coli Ressotta strain. The expression products of rHBcAg(MIR)/MAGE-A3(EPI-1, or -2, or -3) were purified respectively by Ni2+ chelated affinity column, and then confirmed by SDS-PAGE and Western-blot analysis.Purified three rHBcAg(MIR)/MAGE-A3 multiepitopes were administrated respectively into BALB/c (H-2Kd) mice by intradermal injection. The production of rHBcAg(MIR)/MAGE-A3(EPI-1, or -2, or -3) specific IgG in serum from immunized mice were measured by ELISA. Spleen cells from all immunized mice were harvested after one week of last immunization for lymphocyte proliferation assay and cytotoxic T-lymphocyte assay. RESULTS: PCR and Sequencing analysis showed the presence of the required gene fragment in pET21a(+)/ HBcAg(MIR)/MAGE-A3(EPI-1, or -2, or -3) recombinant plasmid. Purified rHBcAg(MIR)/MAGE-A3(EPI-1, or -2, or -3) could be probed specifically by McAb of 6×his-tag. ELISA analysis indicated that serum from immunized mice with rHBcAg(MIR)/MAGE-A3(EPI-1, -2, or -3) proteins could be discerned specifically by complete MAGE-A3 protein, and high level of antibodies in immune serum were obtained, and all antibody titers could reach above 1:1600. The splenocytes from groups of rHBcAg(MIR)/MAGE-A3(EPI-1,-2, or -3), stimulated respectively with corresponding peptides showed the higher proliferative responses comparing with control groups of HBcAg(MIR) or PBS (p<0.05, respectively). Splenocytes from mice immunized with rHBcAg(MIR)/MAGE-A3 (EPI-1, or -2, or -3) could killed target cells effectively, and there were significant difference of CTL activities compared with control groups of HBcAg(MIR), or PBS (p<0.05, respectively) at any ratio of effector : target. CONCLUSION: Our results indicated MIR in HBcAg presenting platform could present MAGE-A3 multiepitopes efficiently and induced significant humoral or cellular immunity. The immune strategy based on multiepitopeimmunization could have potential for preventing or controlling MAGE-A3 associated malignant disease.