Publications by authors named "Gene Kurosawa"

29 Publications

  • Page 1 of 1

β-glucans: wide-spectrum immune-balancing food-supplement-based enteric (β-WIFE) vaccine adjuvant approach to COVID-19.

Hum Vaccin Immunother 2021 Mar 2:1-6. Epub 2021 Mar 2.

The Fujio-Eiji Academic Terrain (FEAT), Nichi-In Centre for Regenerative Medicine (NCRM), Chennai, India.

Conventional vaccines to combat COVID-19 through different approaches are at various stages of development. The complexity of COVID-19 such as the potential mutations of the virus leading to antigenic drift and the uncertainty on the duration of the immunity induced by the vaccine have hampered the efforts to control the COVID-19 pandemic. Thus, we suggest an alternative interim treatment strategy based on biological response modifier glucans such as the AFO-202-derived β-glucan, which has been reported to induce trained immunity, akin to that induced by the Bacille Calmette-Guérin vaccine, by epigenetic modifications at the central level in the bone marrow. These β-glucans act as pathogen-associated molecular patterns, activating mucosal immunity by binding with specific pathogen recognition receptors such as dectin-1 and inducing both the adaptive and innate immunity by reaching distant lymphoid organs. β-Glucans have also been used as immune adjuvants for vaccines such as the influenza vaccine. Therefore, until a conventional vaccine is widely available, an orally consumable vaccine adjuvant that acts like biosimilars, termed as the wide-spectrum immune-balancing food-supplement-based enteric (β-WIFE) vaccine adjuvant approach, with well-reported safety is worth in-depth investigation and can be considered for a clinical trial.
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http://dx.doi.org/10.1080/21645515.2021.1880210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7938654PMC
March 2021

Comprehensive Analysis of Antibodies Induced by Vaccination with 4 Kinds of Avian Influenza H5N1 Pre-Pandemic Vaccines.

Int J Mol Sci 2020 Oct 8;21(19). Epub 2020 Oct 8.

Department of Innovation for Advanced Medicine, Center for Research Promotion and Support, Fujita Health University, Toyoake, Aichi 470-1192, Japan.

Four kinds of avian-derived H5N1 influenza virus, A/Vietnam/1194/2004 (Clade 1), A/Indonesia/5/2005 (Clade 2.1), A/Qinghai/1A/2005 (Clade 2.2), and A/Anhui/1/2005 (Clade 2.3), have been stocked in Japan for use as pre-pandemic vaccines. When a pandemic occurs, these viruses would be used as vaccines in the hope of inducing immunity against the pandemic virus. We analyzed the specificity of antibodies (Abs) produced by B lymphocytes present in the blood after immunization with these vaccines. Eighteen volunteers took part in this project. After libraries of Ab-encoding sequences were constructed using blood from subjects vaccinated with these viruses, a large number of clones that encoded Abs that bound to the virus particles used as vaccines were isolated. These clones were classified into two groups according to the hemagglutination inhibition (HI) activity of the encoded Abs. While two-thirds of the clones were HI positive, the encoded Abs exhibited only restricted strain specificity. On the other hand, half of the HI-negative clones encoded Abs that bound not only to the H5N1 virus but also to the H1N1 virus; with a few exceptions, these Abs appeared to be encoded by memory B cells present before vaccination. The HI-negative clones included those encoding broadly cross-reactive Abs, some of which were encoded by non-V1-69 germline genes. However, although this work shows that various kinds of anti-H5N1 Abs are encoded by volunteers vaccinated with pre-pandemic vaccines, broad cross-reactivity was seen only in a minority of clones, raising concern regarding the utility of these H5N1 vaccine viruses for the prevention of H5N1 pandemics.
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http://dx.doi.org/10.3390/ijms21197422DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7582428PMC
October 2020

Development of anti-human CADM1 monoclonal antibodies as a potential therapy for adult T-cell leukemia/lymphoma.

Int J Hematol 2020 Oct 12;112(4):496-503. Epub 2020 Jul 12.

Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki, 889-1692, Japan.

Adult T-cell leukemia/lymphoma (ATLL) is a highly invasive and refractory T-cell malignancy, with poor prognosis. We previously identified that cell adhesion molecule 1 (CADM1) is overexpressed consistently in ATLL cells, and that CADM1 expression increases the adhesion capacity of ATLL cells to endothelial cells and promotes the organ invasion of ATLL cells in a xenograft mouse model. In this study, we first show that newly developed several anti-human CADM1 antibodies, which were complete human IgG antibodies generated by phage display method, specifically recognize CADM1 on ATLL cells. Although most of the CADM1 antibodies did not have a direct cytotoxic effect against CADM1-positive ATLL cells, clone 089-084 exhibited weak but significant antibody-dependent cell-mediated cytotoxic activity. Moreover, clone 103-189 effectively inhibits the interaction between endothelial cells and CADM1-positive ATLL cells. Furthermore, in mice bearing intra-splenic transplantation of EL4 mouse lymphoma cells expressing CADM1, the treatment of 103-189 significantly suppressed the organ invasion of CADM1-positive EL4 cells, resulting in improved survival time of mice. Therefore, since the anti-CADM1 antibody may be useful for the suppression of organ invasion in ATLL patients, combination use of the anti-CADM1 antibody with chemotherapy drugs could be beneficial for the efficient elimination of ATLL cells.
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http://dx.doi.org/10.1007/s12185-020-02939-1DOI Listing
October 2020

Radiolabeled Human Monoclonal Antibody 067-213 has the Potential for Noninvasive Quantification of CD73 Expression.

Int J Mol Sci 2020 Mar 26;21(7). Epub 2020 Mar 26.

Department of Molecular Imaging and Theranostics, National Institute of Radiological Sciences, National Institutes for Quantum and Radiological Science and Technology (QST-NIRS), Inage, Chiba 263-8555, Japan.

Background: CD73 is an ectonucleotidase regulating extracellular adenosine concentration and plays an important role in adenosine-mediated immunosuppressive pathways. The efficacy of CD73-targeted therapy depends on the expression levels of CD73; therefore, monitoring CD73 status in cancer patients would provide helpful information for selection of patients who would benefit from CD73-targeted therapy. Here, we evaluated the ability of In-labeled antibody 067-213, which has high affinity for human CD73, to act as a noninvasive imaging probe.

Methods: Cell binding and competitive inhibition assays for In-labeled 067-213 were conducted using MIAPaCa-2 (high CD73 expression) and A431 (low CD73 expression) cells. For in vivo assessments, biodistribution and SPECT/CT studies were conducted in MIAPaCa-2 and A431 tumor-bearing mice. To estimate the absorbed dose in humans, biodistribution and SPECT/CT studies were conducted in healthy rats.

Results: In-labeled 067-213 bound to MIAPaCa-2 and A431 cells in a CD73-dependent manner and the affinity loss after In-labeling was limited. Biodistribution and SPECT/CT studies with In-labeled 067-213 in mice showed high uptake in MIAPaCa-2 tumors and lower uptake in A431 tumors. In rats, the probe did not show high uptake in normal organs, including endogenously CD73-expressing organs. The estimated absorbed doses in humans were reasonably low.

Conclusions: In-labeled 067-213 showed CD73-expression-dependent tumor uptake and low uptake in normal organs and tissues. Radiolabeled 067-213 holds promise as an imaging probe for noninvasive evaluation of CD73 expression levels in patients. Our data encourage further clinical studies to clarify a role for CD73 monitoring in patients receiving CD73-targeted immune therapy.
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http://dx.doi.org/10.3390/ijms21072304DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7177856PMC
March 2020

Efficacy Evaluation of Combination Treatment Using Gemcitabine and Radioimmunotherapy with Y-Labeled Fully Human Anti-CD147 Monoclonal Antibody 059-053 in a BxPC-3 Xenograft Mouse Model of Refractory Pancreatic Cancer.

Int J Mol Sci 2018 Sep 29;19(10). Epub 2018 Sep 29.

Department of Molecular Imaging and Theranostics, National Institute of Radiological Sciences, National Institutes for Quantum and Radiological Science and Technology (QST-NIRS), 4-9-1 Anagawa, Inage-ku, Chiba 263-8555, Japan.

The poor prognosis of pancreatic cancer requires the development of more effective therapy. CD147 expresses in pancreatic cancer with high incidence and has a crucial role in invasion and metastasis. We developed a fully human monoclonal antibody (059-053) with high affinity for CD147. Here we evaluated the efficacy of combined treatment using radioimmunotherapy (RIT) with Y-labeled 059-053 and gemcitabine in a BxPC-3 xenograft mouse model. Expression of CD147 and matrix metalloproteinase-2 (MMP2) in BxPC-3 tumors was evaluated. In vitro and in vivo properties of 059-053 were evaluated using In-labeled 059-053 and a pancreatic cancer model BxPC-3. Tumor volume and body weight were periodically measured in mice receiving gemcitabine, RIT, and both RIT and gemcitabine (one cycle and two cycles). High expression of CD147 and MMP2 was observed in BxPC-3 tumors and suppressed by 059-053 injection. Radiolabeled 059-053 bound specifically to BxPC-3 cells and accumulated highly in BxPC-3 tumors but low in major organs. Combined treatment using RIT with gemcitabine (one cycle) significantly suppressed tumor growth and prolonged survival with tolerable toxicity. The two-cycle regimen had the highest anti-tumor effect, but was not tolerable. Combined treatment with Y-labeled 059-053 and gemcitabine is a promising therapeutic option for pancreatic cancer.
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http://dx.doi.org/10.3390/ijms19102979DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6213240PMC
September 2018

A luciferase complementation assay system using transferable mouse artificial chromosomes to monitor protein-protein interactions mediated by G protein-coupled receptors.

Cytotechnology 2018 Dec 31;70(6):1499-1508. Epub 2018 Jul 31.

Chromosome Engineering Research Center, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan.

G protein-coupled receptors (GPCRs) are seven-transmembrane domain receptors that interact with the β-arrestin family, particularly β-arrestin 1 (ARRB1). GPCRs interact with 33% of small molecule drugs. Ligand screening is promising for drug discovery concerning GPCR-related diseases. Luciferase complementation assay (LCA) enables detection of protein-protein complementation via bioluminescence following complementation of N- and C-terminal luciferase fragments (NEluc and CEluc) fused to target proteins, but it is necessary to co-express the two genes. Here, we developed LCAs with mouse artificial chromosomes (MACs) that have unique characteristics such as stable maintenance and a substantial insert-carrying capacity. First, an NEluc-ARRB1 was inserted into MAC4 by Cre-loxP recombination in CHO cells, named ARRB1-MAC4. Second, a parathyroid hormone receptor 2 (PTHR2)-CEluc or prostaglandin EP4 receptor (hEP4)-CEluc were inserted into ARRB1-MAC4, named ARRB1-PTHR2-MAC4 and ARRB1-hEP4-MAC4, respectively, via the sequential integration of multiple vectors (SIM) system. Each MAC was transferred into HEK293 cells by microcell-mediated chromosome transfer (MMCT). LCAs using the established HEK293 cell lines resulted in 35,000 photon counts upon somatostatin stimulation for ARRB1-MAC4 with transient transfection of the somatostatin receptor 2 (SSTR2) expression vector, 1800 photon counts upon parathyroid hormone stimulation for ARRB1-PTHR2-MAC4, and 35,000 photon counts upon prostaglandin E2 stimulation for ARRB1-hEP4-MAC4. These MACs were maintained independently from host chromosomes in CHO and HEK293 cells. HEK293 cells containing ARRB1-PTHR2-MAC4 showed a stable reaction for long-term. Thus, the combination of gene loading by the SIM system into a MAC and an LCA targeting GPCRs provides a novel and useful platform to discover drugs for GPCR-related diseases.
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http://dx.doi.org/10.1007/s10616-018-0231-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6269364PMC
December 2018

Three Types of Broadly Reacting Antibodies against Influenza B Viruses Induced by Vaccination with Seasonal Influenza Viruses.

J Immunol Res 2018 8;2018:7251793. Epub 2018 May 8.

Department of Innovation for Advanced Medicine, Center for Research Promotion and Support, Fujita Health University, Toyoake, Aichi 470-1192, Japan.

We analyzed the antibody (Ab) repertoire against influenza B viruses induced by vaccination with seasonal influenza viruses in one individual who had never been vaccinated until 2009. The vaccine used in this study comprised B/Massachusetts/2/2012 (Yamagata lineage), A/Texas/50/2012 (H3N2), and A/California/7/2009 (H1N1). One month after the subject received two vaccinations, blood (200 ml) was obtained and peripheral mononuclear cells were prepared, and a large Ab library was constructed using phage display technology. The library was screened with HA-enriched fraction of B/Massachusetts/2/2012 and B/Brisbane/60/2008 (Victoria lineage) virus, and a total of 26 Abs that potentially bound to hemagglutinin (HA) molecules were isolated. Their binding activities to six influenza B viruses, three of Yamagata lineage and three of Victoria lineage, and two influenza A viruses, H1N1 and H3N2, were examined. The Abs showed cross-reactivity at three different levels. The first type bound to all Yamagata lineage viruses. The second type bound to both Yamagata and Victoria lineage viruses. The third type bound to both influenza A and B viruses. These results indicate that common epitopes exist on HA molecules of influenza virus at various levels, and humans have capability to produce Abs that bind to such common epitopes.
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http://dx.doi.org/10.1155/2018/7251793DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5964595PMC
October 2018

Development of a complete human IgG monoclonal antibody to transferrin receptor 1 targeted for adult T-cell leukemia/lymphoma.

Biochem Biophys Res Commun 2017 03 8;485(1):144-151. Epub 2017 Feb 8.

Division of Tumor and Cellular Biochemistry, Department of Medical Science, Faculty of Medicine, University of Miyazaki, Japan. Electronic address:

Iron is an essential nutrient for normal cell growth, and reprogramming of iron metabolism is essential to tumor cell survival and progression. HTLV-1-associated adult T-cell leukemia/lymphoma (ATLL) has no effective therapy and high levels of cell surface transferrin receptor 1 (TFR1) expression have been reported in ATLL by us and other groups. In this study, to develop a novel molecular-targeted therapy against TFR1 to modulate iron metabolism, we initially determined the expression pattern of several iron-related genes along with TFR1 and found that ATLL cells presented characteristic of an iron-deficiency state such as high expression of iron-regulatory protein 2 (IRP2) and low expression of its E3 ubiquitin-ligase, FBXL5. Therefore, we developed human IgG monoclonal antibodies to human TFR1 using a phage display method (ICOS method) to block the incorporation of the transferrin (TF)-iron complex into ATLL cells for inhibiting cell growth. One of the mAbs, JST-TFR09, presented its greater affinity to TFR1 on ATLL cells in flow cytometry (FCM) analysis than those of commercially available anti-TFR1 antibodies and identified high expression of TFR1 in most of the acute-type ATLL cells. Moreover, JST-TFR09 could interfere with binding between TFR1 and TF, which resulted in effective blockade of TFR1 internalization and induction of cell apoptosis by the treatment of ATLL cells with JST-TFR09. JST-TFR09 showed dual activities through direct cell cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC), and the treatment of JST-TFR09 significantly suppressed cell growth of ATLL cells with induction of apoptosis in in vitro and in vivo experiments. Thus, JST-TFR09 described here may become a promising therapeutic antibody for the treatment of ATLL.
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http://dx.doi.org/10.1016/j.bbrc.2017.02.039DOI Listing
March 2017

Uptake of 111In-labeled fully human monoclonal antibody TSP-A18 reflects transferrin receptor expression in normal organs and tissues of mice.

Oncol Rep 2017 Mar 30;37(3):1529-1536. Epub 2017 Jan 30.

Department of Molecular Imaging and Theranostics, National Institute of Radiological Sciences, Inage-ku, Chiba 263-8555, Japan.

Transferrin receptor (TfR) is an attractive molecule for targeted therapy of cancer. Various TfR-targeted therapeutic agents such as anti-TfR antibodies conjugated with anticancer agents have been developed. An antibody that recognizes both human and murine TfR is needed to predict the toxicity of antibody-based agents before clinical trials, there is no such antibody to date. In this study, a new fully human monoclonal antibody TSP-A18 that recognizes both human and murine TfR was developed and the correlation analysis of the radiolabeled antibody uptake and TfR expression in two murine strains was conducted. TSP-A18 was selected using extracellular portions of human and murine TfR from a human antibody library. The cross-reactivity of TSP-A18 with human and murine cells was confirmed by flow cytometry. Cell binding and competitive inhibition assays with [111In]TSP-A18 showed that TSP-A18 bound highly to TfR-expressing MIAPaCa-2 cells with high affinity. Biodistribution studies of [111In]TSP-A18 and [67Ga]citrate (a transferrin-mediated imaging probe) were conducted in C57BL/6J and BALB/c-nu/nu mice. [111In]TSP-A18 was accumulated highly in the spleen and bone containing marrow component of both strains, whereas high [67Ga]citrate uptake was only observed in bone containing marrow component and not in the spleen. Western blotting indicated the spleen showed the strongest TfR expression compared with other organs in both strains. There was significant correlation between [111In]TSP-A18 uptake and TfR protein expression in both strains, whereas there was significant correlation of [67Ga]citrate uptake with TfR expression only in C57BL/6J. These findings suggest that the difference in TfR expression between murine strains should be carefully considered when testing for the toxicity of anti-TfR antibody in mice and the uptake of anti-TfR antibody could reflect tissue TfR expression more accurately compared with that of transferrin-mediated imaging probe such as [67Ga]citrate.
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http://dx.doi.org/10.3892/or.2017.5412DOI Listing
March 2017

Classification of 27 Tumor-Associated Antigens by Histochemical Analysis of 36 Freshly Resected Lung Cancer Tissues.

Int J Mol Sci 2016 Nov 8;17(11). Epub 2016 Nov 8.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake 470-1192, Japan.

In previous studies, we identified 29 tumor-associated antigens (TAAs) and isolated 488 human monoclonal antibodies (mAbs) that specifically bind to one of the 29 TAAs. In the present study, we performed histochemical analysis of 36 freshly resected lung cancer tissues by using 60 mAbs against 27 TAAs. Comparison of the staining patterns of tumor cells, bronchial epithelial cells, and normal pulmonary alveolus cells and interalveolar septum allowed us to determine the type and location of cells that express target molecules, as well as the degree of expression. The patterns were classified into 7 categories. While multiple Abs were used against certain TAAs, the differences observed among them should be derived from differences in the binding activity and/or the epitope. Thus, such data indicate the versatility of respective clones as anti-cancer drugs. Although the information obtained was limited to the lung and bronchial tube, bronchial epithelial cells represent normal growing cells, and therefore, the data are informative. The results indicate that 9 of the 27 TAAs are suitable targets for therapeutic Abs. These 9 Ags include EGFR, HER2, TfR, and integrin α6β4. Based on our findings, a pharmaceutical company has started to develop anti-cancer drugs by using Abs to TfR and integrin α6β4. HGFR, PTP-LAR, CD147, CDCP1, and integrin αvβ3 are also appropriate targets for therapeutic purposes.
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http://dx.doi.org/10.3390/ijms17111862DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5133862PMC
November 2016

A variety of human monoclonal antibodies against epidermal growth factor receptor isolated from a phage antibody library.

Biochem Biophys Res Commun 2016 Nov 5;480(1):94-100. Epub 2016 Oct 5.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, Japan. Electronic address:

When the technology for constructing human antibody (Ab) libraries using a phage-display system was developed, many researchers in Ab-related fields anticipated that it would be widely applied to the development of pharmaceutical drugs against various diseases, including cancers. However, successful examples of such applications are very limited. Moreover, researchers who utilize phage-display technology now show divergent ways of thinking about phage Ab libraries. For example, there is debate about what should be the source of V and V genes for the construction of libraries to cover the whole repertoire of Abs present in the human body. In the immune system, the introduction of mutations into V genes followed by selection based on binding activity, termed Ab maturation, is required for the production of Abs exhibiting high affinity to the antigen (Ag). Therefore, introduction of mutations and selection are required for isolation of Abs with high affinity after isolation of clones from phage Ab libraries. We constructed a large human Ab library termed AIMS, developed a screening method termed ICOS, and succeeded in isolating many human monoclonal Abs (mAbs) that specifically and strongly bind to various tumor-associated Ags. Eight anti-EGFR mAbs were included, which we characterized. These mAbs showed various different activities against EGFR-expressing cancer cells. In this paper, we describe these data and discuss the possibility and necessity that the mAbs isolated from the AIMS library might be developed as therapeutic drugs against cancers without introduction of mutations.
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http://dx.doi.org/10.1016/j.bbrc.2016.10.002DOI Listing
November 2016

Trauma team discord and the role of briefing.

J Trauma Acute Care Surg 2016 07;81(1):184-9

From The Queen's Medical Center (S.S., A.B., G.S.); Department of Surgery (S.S.), Division of Emergency Medicine (A.B., G.S.), Biostatistics and Quantitative Health Sciences (E.L.), SimTiki Simulation Center (B.B.), University of Hawaii, John A. Burns School of Medicine (A.W., N.H., G.K.).

Background: Briefing of the trauma team before patient arrival is unstructured in many centers. We surveyed trauma teams regarding agreement on patient care priorities and evaluated the impact of a structured, physician-led briefing on concordance during simulated resuscitations.

Methods: Trauma nurses at our Level II center were surveyed, and they participated in four resuscitation scenarios, randomized to "briefed" or "nonbriefed." For nonbriefed scenarios, nurses independently reviewed triage sheets with written information. Briefed scenarios had a structured 4-minute physician-led briefing reviewing triage sheets identical to nonbriefed scenarios. Teams included three to four nurses (subjects) and two to four confederates (physicians, respiratory therapists). Each team served as their own control group. Confederates were blinded to nurses' briefed or nonbriefed status. Immediately before, and at the midpoint of each scenario, nurses estimated patients' morbidity and mortality and ranked the top 3 of 16 designated immediate care priorities. Briefed and nonbriefed groups' responses were compared for (1) agreement using intraclass correlation coefficient, (2) concordance with physicians' responses using the Fisher exact test, (3) teamwork via T-NOTECHS ratings by nurses and physicians using t-test, and (4) time to complete clinical tasks using t test.

Results: Thirty-eight nurses participated. Ninety-seven percent "agreed/strongly agreed" briefing is important, but only 46% agreed briefing was done well. Comparing briefed versus nonbriefed scenarios, nurses' estimation of morbidity and mortality in the briefed scenarios showed significantly greater agreement with each other and with physicians' answers (p < 0.01). Rank lists also better agreed with each other (intraclass correlation coefficient, 0.64 vs 0.59) and with physicians' answers in the briefed scenarios. T-NOTECHS Leadership ratings were significantly higher in the briefed scenarios (3.70 vs 3.39; p < 0.01). Time to completion of key clinical tasks was significantly faster for one of the briefed scenarios.

Conclusions: Discordant perceptions of patient care goals was frequently observed. Structured physician-led briefing seemed to improve interprofessional team concordance, leadership, and task completion in simulated trauma resuscitations.
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http://dx.doi.org/10.1097/TA.0000000000001024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4915979PMC
July 2016

Role confusion and self-assessment in interprofessional trauma teams.

Am J Surg 2016 Feb 12;211(2):482-8. Epub 2015 Dec 12.

Department of Medicine, John A. Burns School of Medicine, University of Hawaii, Honolulu, HI.

Background: Trauma care requires coordinating an interprofessional team, with formative feedback on teamwork skills. We hypothesized nurses and surgeons have different perceptions regarding roles during resuscitation; that nurses' teamwork self-assessment differs from experts', and that video debriefing might improve accuracy of self-assessment.

Methods: Trauma nurses and surgeons were surveyed regarding resuscitation responsibilities. Subsequently, nurses joined interprofessional teams in simulated trauma resuscitations. After each resuscitation, nurses and teamwork experts independently scored teamwork (T-NOTECHS). After video debriefing, nurses repeated T-NOTECHS self-assessment.

Results: Nurses and surgeons assumed significantly more responsibility by their own profession for 71% of resuscitation tasks. Nurses' overall T-NOTECHS ratings were slightly higher than experts'. This was evident in all T-NOTECHS subdomains except "leadership," but despite statistical significance the difference was small and clinically irrelevant. Video debriefing did not improve the accuracy of self-assessment.

Conclusions: Nurses and physicians demonstrated discordant perceptions of responsibilities. Nurses' self-assessment of teamwork was statistically, but not clinically significantly, higher than experts' in all domains except physician leadership.
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http://dx.doi.org/10.1016/j.amjsurg.2015.11.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4724377PMC
February 2016

Evaluation of Efficacy of Radioimmunotherapy with 90Y-Labeled Fully Human Anti-Transferrin Receptor Monoclonal Antibody in Pancreatic Cancer Mouse Models.

PLoS One 2015 20;10(4):e0123761. Epub 2015 Apr 20.

Diagnostic Imaging Program, Molecular Imaging Center, National Institute of Radiological Sciences, Inage-ku, Chiba, Japan.

Objective: Pancreatic cancer is an aggressive tumor and the prognosis remains poor. Therefore, development of more effective therapy is needed. We previously reported that 89Zr-labeled TSP-A01, an antibody against transferrin receptor (TfR), is highly accumulated in a pancreatic cancer xenograft, but not in major normal organs. In the present study, we evaluated the efficacy of radioimmunotherapy (RIT) with 90Y-TSP-A01 in pancreatic cancer mouse models.

Methods: TfR expression in pancreatic cancer cell lines (AsPC-1, BxPC-3, MIAPaCa-2) was evaluated by immunofluorescence staining. 111In-labeled anti-TfR antibodies (TSP-A01, TSP-A02) were evaluated in vitro by cell binding assay with the three cell lines and by competitive inhibition assay with MIAPaCa-2. In vivo biodistribution was evaluated in mice bearing BxPC-3 and MIAPaCa-2 xenografts. Tumor volumes of BxPC-3 and MIAPaCa-2 were sequentially measured after 90Y-TSP-A01 injection and histological analysis of tumors was conducted.

Results: MIAPaCa-2 cells showed the highest TfR expression, followed by AsPC-1 and BxPC-3 cells. 111In-TSP-A01 and 111In-TSP-A02 bound specifically to the three cell lines according to TfR expression. The dissociation constants for TSP-A01, DOTA-TSP-A01, TSP-A02, and DOTA-TSP-A02 were 0.22, 0.28, 0.17, and 0.22 nM, respectively. 111In-TSP-A01 was highly accumulated in tumors, especially in MIAPaCa-2, but this was not true of 111In-TSP-A02. The absorbed dose for 90Y-TSP-A01 was estimated to be 8.3 Gy/MBq to BxPC-3 and 12.4 Gy/MBq to MIAPaCa-2. MIAPaCa-2 tumors treated with 3.7 MBq of 90Y-TSP-A01 had almost completely disappeared around 3 weeks after injection and regrowth was not observed. Growth of BxPC-3 tumors was inhibited by 3.7 MBq of 90Y-TSP-A01, but the tumor size was not reduced.

Conclusion: 90Y-TSP-A01 treatment achieved an almost complete response in MIAPaCa-2 tumors, whereas it merely inhibited the growth of BxPC-3 tumors. 90Y-TSP-A01 is a promising RIT agent for pancreatic cancer, although further investigation is necessary to improve the efficacy for the radioresistant types like BxPC-3.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0123761PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4404254PMC
January 2016

Preclinical evaluation of ⁸⁹Zr-labeled human antitransferrin receptor monoclonal antibody as a PET probe using a pancreatic cancer mouse model.

Nucl Med Commun 2015 Mar;36(3):286-94

aDiagnostic Imaging Program bMolecular Probe Program, Molecular Imaging Center, National Institute of Radiological Sciences, Chiba cDepartment of Nuclear Medicine, Cancer Institute Hospital dResearch and Development Division, Perseus Proteomics Inc., Tokyo eInnovation Center for Advanced Medicine, School of Medicine, Fujita Health University, Aichi, Japan.

Objective: Pancreatic cancer is aggressive and its prognosis remains poor; thus, effective therapy is urgently needed. Transferrin receptor (TfR) is highly expressed in pancreatic cancer and is considered to be a good candidate for molecular-targeted therapy. We radiolabeled and evaluated fully human anti-TfR monoclonal antibodies as a new PET probe for evaluating the biodistribution of the anti-TfR antibody in pancreatic cancer.

Materials And Methods: TfR expression was evaluated in four human pancreatic cancer (MIAPaCa-2, PANC-1, BxPC-3, and AsPC-1) and murine A4 cell lines. The binding of 125I-labeled anti-TfR antibodies (TSP-A01, TSP-A02, TSP-A03, and TSP-A04) to MIAPaCa-2 cells was compared. 125I-labeled, 67Ga-labeled, and 89Zr-labeled TSP-A01 were evaluated by cell binding, competitive inhibition, and internalization assays. Biodistribution studies of 125I-labeled and 89Zr-labeled TSP-A01 were conducted in mice bearing MIAPaCa-2 and A4 tumors. PET imaging with [89Zr]TSP-A01 was carried out.

Results: MIAPaCa-2 cells showed the highest TfR expression in vitro and in vivo, whereas A4 cells showed no expression. Of the four antibodies, [125I]TSP-A01 showed the highest binding to MIAPaCa-2 cells, but not to A4 cells. The dissociation constant of TSP-A01 was 0.29 nmol/l. Uptake of radiolabeled TSP-A01, especially [89Zr]TSP-A01, was significantly higher in MIAPaCa-2 tumors than in A4 tumors. PET with [89Zr]TSP-A01 clearly visualized MIAPaCa-2 xenografts but not A4 xenografts.

Conclusion: [89Zr]TSP-A01 is a promising PET probe for evaluating the accumulation of anti-TfR antibody in pancreatic cancer and has the potential to facilitate the selection of appropriate patients who would benefit from anti-TfR antibody therapy.
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http://dx.doi.org/10.1097/MNM.0000000000000245DOI Listing
March 2015

Development of a complete human anti-human transferrin receptor C antibody as a novel marker of oral dysplasia and oral cancer.

Cancer Med 2014 Aug 2;3(4):1085-99. Epub 2014 Jun 2.

Division of Oral and Maxillofacial Surgery, Medicine of Sensory and Motor Organs, University of Miyazaki, Miyazaki, Japan; Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

Oral squamous cell carcinoma (OSCC) is the sixth most common cancer worldwide. Up to 20% of oral dysplasia cases have been suggested to undergo malignant transformation to OSCC; however, there are no methods to predict OSCC development. In this study, to identify the genes associated with oral dysplasia progression, we performed genomic copy number analyses of genomic DNA samples isolated from primary oral dysplasia and OSCC via the microdissection method and found elevated expression of transferrin receptor C (TfR1/TFRC) with genomic amplification in oral dysplasia and OSCC. The expression rate of TFRC in OSCC was significantly higher than that in dysplasia, suggesting that OSCC disease progression might be related to TFRC expression. Additionally, we investigated the in vitro and in vivo impacts of a newly established anti-human TFRC monoclonal antibody, which was isolated from a human cDNA library using the phage-display method, on cell proliferation and survival. The anti-TFRC antibody blocked the interaction between transferrin and TFRC and consequently inhibited iron uptake, leading to the iron deprivation-mediated suppression of cell growth and induction of apoptosis. Moreover, we demonstrated that the anti-TFRC antibody efficiently inhibited tumor growth in a murine xenograft OSCC model. Therefore, we suggest our developed complete human anti-human TFRC antibody as a useful, novel treatment for oral dysplasia and OSCC.
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http://dx.doi.org/10.1002/cam4.267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4303177PMC
August 2014

Smallpox still haunts scientists: results of a questionnaire-based inquiry on the views of health care and life science experts and students on preserving the remaining variola virus stocks.

ScientificWorldJournal 2013 22;2013:672813. Epub 2013 Jul 22.

The Fujio-Eiji Academic Terrain (FEAT), Nichi-In Centre for Regenerative Medicine (NCRM), Chennai 600034, India.

The World Health Organization (WHO) declared eradication of the dreadful disease "smallpox" in 1980. Though the disease has died down, the causative virus "variola" has not, as it has been well preserved in two high security laboratories-one in USA and another in Russia. The debate on whether the remaining stocks of the smallpox virus should be destroyed or not is ongoing, and the World Health Assembly (WHA) in 2011 has decided to postpone the review on this debate to the 67th WHA in 2014. A short questionnaire-based inquiry was organized during a one-day stem cell meeting to explore the views of various health care and life science specialists especially students on this aspect. Among the 200 participants of the meeting, only 66 had answered the questionnaire. 60.6% of participants who responded to the questionnaire were for preserving the virus for future reference, while 36.4% of the participants were for destroying the virus considering the magnitude with which it killed millions. However, 3% of the respondents were not able to decide on any verdict. Therefore, this inquiry expresses the view that "what we cannot create, we do not have the right to destroy."
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http://dx.doi.org/10.1155/2013/672813DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3736420PMC
February 2014

Evaluation of (89)Zr-labeled human anti-CD147 monoclonal antibody as a positron emission tomography probe in a mouse model of pancreatic cancer.

PLoS One 2013 5;8(4):e61230. Epub 2013 Apr 5.

Diagnostic Imaging Group, Molecular Imaging Center, National Institute of Radiological Sciences, Chiba, Japan.

Introduction: Pancreatic cancer is an aggressive cancer and its prognosis remains poor. Therefore, additional effective therapy is required to augment and/or complement current therapy. CD147, high expression in pancreatic cancer, is involved in the metastatic process and is considered a good candidate for targeted therapy. CD147-specfic imaging could be useful for selection of appropriate patients. Therefore, we evaluated the potential of a fully human anti-CD147 monoclonal antibody 059-053 as a new positron emission tomography (PET) probe for pancreatic cancer.

Methods: CD147 expression was evaluated in four pancreatic cancer cell lines (MIA Paca-2, PANC-1, BxPC-3, and AsPC-1) and a mouse cell line A4 as a negative control. Cell binding, competitive inhibition and internalization assays were conducted with (125)I-, (67)Ga-, or (89)Zr-labeled 059-053. In vivo biodistribution of (125)I- or (89)Zr-labeled 059-053 was conducted in mice bearing MIA Paca-2 and A4 tumors. PET imaging with [(89)Zr]059-053 was conducted in subcutaneous and orthotopic tumor mouse models.

Results: Among four pancreatic cancer cell lines, MIA Paca-2 cells showed the highest expression of CD147, while A4 cells had no expression. Immunohistochemical staining showed that MIA Paca-2 xenografts also highly expressed CD147 in vivo. Radiolabeled 059-053 specifically bound to MIA Paca-2 cells with high affinity, but not to A4. [(89)Zr]059-053 uptake in MIA Paca-2 tumors increased with time from 11.0±1.3% injected dose per gram (ID/g) at day 1 to 16.9±3.2% ID/g at day 6, while [(125)I]059-053 uptake was relatively low and decreased with time, suggesting that 059-053 was internalized into tumor cells in vivo and (125)I was released from the cells. PET with [(89)Zr]059-053 clearly visualized subcutaneous and orthotopic tumors.

Conclusion: [(89)Zr]059-053 is a promising PET probe for imaging CD147 expression in pancreatic cancer and has the potential to select appropriate patients with CD147-expressing tumors who could gain benefit from anti-CD147 therapy.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0061230PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3618331PMC
October 2013

Combining phenotypic and proteomic approaches to identify membrane targets in a 'triple negative' breast cancer cell type.

Mol Cancer 2013 Feb 13;12:11. Epub 2013 Feb 13.

MedImmune, Granta Park, Cambridge, CB21 6GH, UK.

Background: The continued discovery of therapeutic antibodies, which address unmet medical needs, requires the continued discovery of tractable antibody targets. Multiple protein-level target discovery approaches are available and these can be used in combination to extensively survey relevant cell membranomes. In this study, the MDA-MB-231 cell line was selected for membranome survey as it is a 'triple negative' breast cancer cell line, which represents a cancer subtype that is aggressive and has few treatment options.

Methods: The MDA-MB-231 breast carcinoma cell line was used to explore three membranome target discovery approaches, which were used in parallel to cross-validate the significance of identified antigens. A proteomic approach, which used membrane protein enrichment followed by protein identification by mass spectrometry, was used alongside two phenotypic antibody screening approaches. The first phenotypic screening approach was based on hybridoma technology and the second was based on phage display technology. Antibodies isolated by the phenotypic approaches were tested for cell specificity as well as internalisation and the targets identified were compared to each other as well as those identified by the proteomic approach. An anti-CD73 antibody derived from the phage display-based phenotypic approach was tested for binding to other 'triple negative' breast cancer cell lines and tested for tumour growth inhibitory activity in a MDA-MB-231 xenograft model.

Results: All of the approaches identified multiple cell surface markers, including integrins, CD44, EGFR, CD71, galectin-3, CD73 and BCAM, some of which had been previously confirmed as being tractable to antibody therapy. In total, 40 cell surface markers were identified for further study. In addition to cell surface marker identification, the phenotypic antibody screening approaches provided reagent antibodies for target validation studies. This is illustrated using the anti-CD73 antibody, which bound other 'triple negative' breast cancer cell lines and produced significant tumour growth inhibitory activity in a MDA-MB-231 xenograft model.

Conclusions: This study has demonstrated that multiple methods are required to successfully analyse the membranome of a desired cell type. It has also successfully demonstrated that phenotypic antibody screening provides a mechanism for rapidly discovering and evaluating antibody tractable targets, which can significantly accelerate the therapeutic discovery process.
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http://dx.doi.org/10.1186/1476-4598-12-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3582597PMC
February 2013

Expression of LY6D is induced at the surface of MCF10A cells by X-ray irradiation.

FEBS J 2012 Dec 22;279(24):4479-91. Epub 2012 Nov 22.

Department of Biochemistry, Nagoya University School of Medicine, Japan.

In order to identify membrane proteins whose expression is induced by X-ray irradiation, we developed an antibody (Ab)-directed strategy using a phage Ab library. X-Ray-irradiated cells were screened with a phage Ab library in the presence of a large excess of polyclonal Abs prepared against membrane proteins that are commonly present at the surface of both X-ray-irradiated and nonirradiated cells. After isolation of Ab that bound only to X-ray-irradiated cells, the antigen was identified using MS. Using this approach, we found that expression of LY6D is induced in MCF10A cells by X-ray irradiation. The induction of LY6D expression is triggered through a pathway regulated by ATM, CHK2 and p53. This method is a new Ab-directed proteomic strategy for analysis of membrane proteins, and is applicable to various biological phenomena in situations in which both target molecule-expressing cells and nonexpressing cells are available.
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http://dx.doi.org/10.1111/febs.12034DOI Listing
December 2012

Novel human monoclonal antibody against epidermal growth factor receptor as an imaging probe for hepatocellular carcinoma.

Nucl Med Commun 2012 Jul;33(7):719-25

Diagnostic Imaging Group, Molecular Imaging Center, National Institute of Radiological Sciences, Chiba, Japan.

Objective: The epidermal growth factor receptor (EGFR) is overexpressed in many epithelial cancers, including hepatocellular carcinoma (HCC), and is an attractive target for cancer imaging and therapy. We attempted a novel noninvasive imaging method to evaluate anti-EGFR human monoclonal antibody clones for determining the uptake of therapeutic anti-EGFR antibody in HCC.

Methods: In-vitro cell binding of nine I-labeled antibody clones was compared in the human epidermoid cancer cell line A431, in three HCC cell lines Hep-G2, SK-Hep1, and HuH-7, and in the EGFR-negative control cell line A4. In-labeled or I-labeled 048-006 was subjected to cell binding, competitive inhibition, and internalization assays using A431, SK-Hep1, and HuH-7. Further, In-labeled 048-006 was evaluated in in-vivo biodistribution analysis and single-photon imaging in nude tumor-bearing mice.

Results: The 048-006 clone showed the highest binding to EGFR-expressing cells among the nine antibodies. In-labeled or I-labeled 048-006 specifically bound to EGFR-expressing cells with high affinity and was internalized after binding to EGFR. A431 and HuH-7 tumors showed high In-labeled 048-006 uptake, which was visualized by single-photon imaging.

Conclusion: Radiolabeled human anti-EGFR monoclonal antibody 048-006 has the potential to be a safer imaging probe for predicting tumor uptake of anti-EGFR antibody therapeutic agents in HCC.
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http://dx.doi.org/10.1097/MNM.0b013e3283531d68DOI Listing
July 2012

Selection and analysis of anti-cancer antibodies for cancer therapy obtained from antibody phage library.

Cancer Sci 2011 Jan 12;102(1):175-81. Epub 2010 Oct 12.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, Japan.

The search for effective antibodies (Ab) for curable cancer immunotherapy has been a quest of many research groups in order to find an effective target that exists on the cancer cell surface. So far there have been no conclusive answers to shed light on the search. This study therefore aimed to bridge the gap of cancer therapy. Screening against 49 kinds of cell lines belonging to 11 kinds of solids cancers was performed. Isolation and characterization for approximately 4200 monoclonal antibodies (mAb) was also performed thereafter. Of those mAb 488 clones that turned out to bind to 29 tumor-associated antigens (TAA) were subjected to immunohistochemical (IHC) analyses. Selection of target antigens (Ag) and a potential antibody for cancer therapy was conducted prior to clinical examinations. In order to find predictably effective targets for therapeutic Ab against solid cancers, expression of the Ag on the surface of cancer and normal cells was extensively examined by IHC analyses using fresh cancer specimens resected from patients. In this study, the tendencies of all staining patterns and distribution of the Ab are reported. While all of the TAA appeared to be involved in tumorigenesis, their expression was not restricted to some specific tumor types but rather randomly distributed among various cancers. Some kinds of Ab including anti-epidermal growth factor receptor (EGFR) and anti-human epidermal growth factor receptor 2 (HER2) indicated the frequency of expression in normal cells was generally low. We concluded that identification of 488 mAb and the accumulated results of IHC analyses in this study could be the key for further therapeutic Ab against cancers. The targets that showed cancer-specific expression are expected to be better for therapeutic Ab than the other Ab. Moreover, further investigation into the growth of cancer cell lines using full human IgG form of Ab shows available efficacy in specific cases.
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http://dx.doi.org/10.1111/j.1349-7006.2010.01739.xDOI Listing
January 2011

Methods for comprehensive identification of membrane proteins recognized by a large number of monoclonal antibodies.

J Immunol Methods 2009 Dec 18;351(1-2):1-12. Epub 2009 Sep 18.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi 470-1192, Japan.

In order to isolate monoclonal antibodies (mAbs) that bind to tumor-associated antigens (Ags) we developed the following strategy. Using the phage-display Ab library we isolated a large number of mAbs that bind to the surface of human tumor cells. The mAbs were individually screened by immunostaining, and clones that preferentially and strongly stained the malignant cells were chosen. Thereafter, the Ags recognized by the mAbs were identified. For identification of the Ags by MS candidate molecules had to be purified either by immunoprecipitation or by affinity chromatography. We isolated several hundred mAbs that showed cancer-specific staining patterns. In order to identify the Ags that were recognized by the numerous mAbs within a short time we developed two methods. Using the GFC [grouping of clones by flow cytometry (FCM)] method many Abs could be grouped by comparing the staining patterns of FCM. Members in each group turned out to bind to the same molecule in many cases. After a candidate Ag was revealed, the polypeptide corresponding to its extracellular portion was prepared and used for identification of clones that bound to the Ag among all the mAbs by SITE (simultaneous identification of clones through three dimensional ELISA) method. Both methods can be generally applicable to various kinds of membrane proteins and the mAbs against them.
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http://dx.doi.org/10.1016/j.jim.2009.09.003DOI Listing
December 2009

Frequent overexpression of CADM1/IGSF4 in lung adenocarcinoma.

Biochem Biophys Res Commun 2009 Jun 14;383(4):480-4. Epub 2009 Apr 14.

Department of Surgery, School of Medicine, Fujita Health University, Toyoake, Aichi 470-1192, Japan.

We reported comprehensive screening for antigens (Ags) overexpressed on various carcinomas via isolation of human monoclonal antibodies (mAbs) that may be therapeutic in a previous paper (Proc. Natl. Acad. Sci. USA 105, 7287-7292, 2008). Twenty-one distinct Ags highly expressed on several carcinomas were identified and 356 mAbs with unique sequences turned out to bind to one of the 21 Ags. Among them CADM1/IGSF4 which had been originally referred to as tumor suppressor lung cancer 1 (TSLC1) was included. Therefore we examined the expression of CADM1 in lung cancers in this study. Eight different anti CADM1 mAbs were used for immunohistochemical analysis of 29 fresh lung cancer specimens. Staining patterns were categorized to six groups based on the extent of positive staining and the localization of stained portions. While overexpression of CADM1 was observed on the cell surface of adenocarcinomas at a high frequency, around 60%, positive stainings were rarely observed on that of other lung carcinomas including squamous cell carcinomas. Moreover, some clones among the eight mAbs gave different staining patterns from those by the other clones against the same fresh specimen, suggesting presence of variant forms of CADM1 differentiated by mAbs.
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http://dx.doi.org/10.1016/j.bbrc.2009.04.039DOI Listing
June 2009

Isolation of antigen/antibody complexes through organic solvent (ICOS) method.

Biochem Biophys Res Commun 2009 Jan 9;378(4):832-5. Epub 2008 Dec 9.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, Japan.

We developed a method termed ICOS (isolation of antigen-antibody complexes through organic solvent) for comprehensive isolation of monoclonal antibodies (mAbs) bound to molecules on the cell surface. By mixing a large number of phage particles of an antibody (Ab) library with living cells, antigen (Ag)-Ab complexes were formed on the cell surface. The mixture was overlaid on organic solution in a tube and subjected to centrifugation. Phages bound to cells were recovered from the precipitate. The phage fraction isolated turned out to contain mAbs that bind to very heterogeneous epitopes and show strong binding activity to Ags. The ICOS method was applied to isolation of human mAbs that may be therapeutic against cancers. Sixty percent of clones isolated by the screening of a phage Ab library against cancer cells turned out to bind to various kinds of tumor-associated Ags. The precise protocol of ICOS method and the rationale of efficient screening were described.
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http://dx.doi.org/10.1016/j.bbrc.2008.11.129DOI Listing
January 2009

Comprehensive screening for antigens overexpressed on carcinomas via isolation of human mAbs that may be therapeutic.

Proc Natl Acad Sci U S A 2008 May 12;105(20):7287-92. Epub 2008 May 12.

Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi 470-1192, Japan.

Although several murine mAbs that have been humanized became useful therapeutic agents against a few malignancies, therapeutic Abs are not yet available for the majority of the human cancers because of our lack of knowledge of which antigens (Ags) can become useful targets. In the present study we established a procedure for comprehensive identification of such Ags through the extensive isolation of human mAbs that may become therapeutic. Using the phage-display Ab library we isolated a large number of human mAbs that bind to the surface of tumor cells. They were individually screened by immunostaining, and clones that preferentially and strongly stained the malignant cells were chosen. The Ags recognized by those clones were isolated by immunoprecipitation and identified by MS. We isolated 2,114 mAbs with unique sequences and identified 21 distinct Ags highly expressed on several carcinomas. Of those 2,114 mAbs 356 bound specifically to one of the 21 Ags. After preparing complete IgG(1) Abs the in vitro assay for Ab-dependent cell-mediated cytotoxicity (ADCC) and the in vivo assay in cancer-bearing athymic mice were performed to examine antitumor activity. The mAbs converted to IgG(1) revealed effective ADCC as well as antitumor activity in vivo. Because half of the 21 Ags showed distinct tumor-specific expression pattern and the mAbs isolated showed various characteristics with strong affinity to the Ag, it is likely that some of the Ags detected will become useful targets for the corresponding carcinoma therapy and that several mAbs will become therapeutic agents.
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http://dx.doi.org/10.1073/pnas.0712202105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2377314PMC
May 2008

Duplicated Abd-B class genes in medaka hoxAa and hoxAb clusters exhibit differential expression patterns in pectoral fin buds.

Dev Genes Evol 2007 Apr 27;217(4):263-73. Epub 2007 Feb 27.

Division of Biological Science, Graduate School of Science, Nagoya University, Nagoya, Aichi 464-8602, Japan.

Hox genes form clusters. Invertebrates and Amphioxus have only one hox cluster, but in vertebrates, they are multiple, i.e., four in the basal teleost fish Polyodon and tetrapods (HoxA, B, C, D), but seven or eight in common teleosts. We earlier completely sequenced the entire hox gene loci in medaka fish, showing a total of 46 hox genes to be encoded in seven clusters (hoxAa, Ab, Ba, Bb, Ca, Da, Db). Among them, hoxAa, hoxAb and hoxDa clusters are presumed to be important for fin-to-limb evolution because of their key role in forelimb and pectoral fin development. In the present study, we compared genome organization and nucleotide sequences of the hoxAa and hoxAb clusters to these of tetrapod HoxA clusters, and found greater similarity in hoxAa case. We then analyzed expression of Abd-B family genes in the clusters. In the trunk, those from the hoxAa cluster, i.e., hoxA9a, hoxA10a, hoxA11a and hoxA13a, were expressed in a manner keeping the colinearity rule of the hox expression as those of tetrapods, while those from the hoxAb cluster, i.e., hoxA9b, hoxA10b, hoxA11b and hoxA13b, were not. In the pectoral fins, the hoxAa cluster was expressed in split domains and did not obey the rule. By contrast, those from the hoxAb and hoxDa clusters were expressed in a manner keeping the rule, i.e., an ancestral pattern similar to those of tetrapods. It is plausible that this differential expression of the two clusters is caused by changes occurred in global control regions after cluster duplications.
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http://dx.doi.org/10.1007/s00427-007-0137-4DOI Listing
April 2007

Organization and structure of hox gene loci in medaka genome and comparison with those of pufferfish and zebrafish genomes.

Gene 2006 Mar 9;370:75-82. Epub 2006 Feb 9.

Division of Biological Sciences, Graduate School of Science, Nagoya University, Nagoya, Aichi 460-8602, Japan.

We isolated BAC clones that cover the entire hox gene loci in the medaka fish Oryzias latipes. The BAC clones were characterized by the Southern hybridization with many hox gene probes isolated in our previous study and by PCR using primers designed for selective amplification of respective hox genes. Then, the BAC clones have been subjected to shotgun sequencing. The results revealed the organization of the entire hox gene loci. Forty-six hox genes in total are encoded in seven clusters as follows: 10 hox genes in Aa cluster; 5 in Ab; 9 in Ba; 4 in Bb; 10 in Ca; 6 in Da; and 2 in Db. Together with the information on the hox gene loci registered in the Fugu genome database and in the Danio genome database, the physical maps of three fish genomes were constructed and compared one another. Not only numbers of hox genes but also the distances between the neighboring hox genes are highly similar between medaka and fugu. As for six clusters, Aa, Ab, Ba, Bb, Ca and Da that are commonly present in the three fishes, only few or no differences were found in each cluster. Thus, the hox gene sets should have been well conserved once they had been established in respective species.
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http://dx.doi.org/10.1016/j.gene.2005.11.015DOI Listing
March 2006