RSSKRAS genotyping in rectal adenocarcinoma specimens with low tumor cellularity after neoadjuvant treatment.
Source
Department of Pathology, Val d'Aurelle Cancer Institute, Montpellier, France.
Abstract
KRAS status assessment is mandatory in patients with metastatic colorectal cancer before therapy with anti-epidermal growth factor receptor monoclonal antibodies, as KRAS mutations are associated with resistance to this treatment. However, KRAS genotyping may be very challenging in case of poor tumor cellularity, particularly when major tumor regression is achieved in locally advanced rectal adenocarcinomas after radiochemotherapy. We aimed at identifying the most reliable strategy to detect KRAS mutations in such samples. DNA was extracted from 31 surgical specimens with major tumor regression, following manual dissection, and from paired pre-treatment biopsies and analyzed by high-resolution melting. DNA samples displaying altered melting curve shapes were then sequenced. Samples with unmodified melting curves or wild-type sequence were further investigated by using an allele-specific PCR assay (TheraScreen) and laser microdissection (followed by high-resolution melting and sequencing analyses). In the 31 post-radiochemotherapy surgical specimens, seven KRAS mutations were identified by high-resolution melting analysis/sequencing. One additional mutation was detected by the TheraScreen assay and two mutations, including the one identified by the TheraScreen assay, were detected following laser microdissection. Altogether, 9/31 surgical specimens (29%) presented KRAS mutations. In the manually dissected pre-treatment biopsies, 12 mutations (39%) were identified by high-resolution melting analysis and sequencing. No additional mutations were found by using the TheraScreen assay or laser microdissection. These results indicate that, in the case of post-radiochemotherapy surgical specimens of colorectal cancer with low tumor cellularity, pre-treatment biopsies might represent the most cost-effective option for reliable KRAS genotyping. The use of more sensitive assays, such as allele-specific PCR or laser microdissection, can be envisaged but with higher costs and longer delays.Modern Pathology advance online publication, 27 January 2012; doi:10.1038/modpathol.2011.210.
- PMID:
- 22282307
- [PubMed - as supplied by publisher]
Discovery and development of N-cadherin antagonists.
Source
Division of Urology, Department of Surgery, McGill University, Montreal, QC, Canada, orest.blaschuk@mcgill.ca.
Abstract
This article describes over 20 years of research on antagonists of the cell adhesion molecule, N-cadherin. Four types of antagonists are discussed: synthetic linear peptides, synthetic cyclic peptides, non-peptidyl peptidomimetics of the disulfide linked cyclic peptide N-Ac-CHAVC-NH(2) and monoclonal antibodies directed against the N-cadherin ectodomain. The biological activities of these antagonists are also discussed. In particular, the ability of N-cadherin antagonists to act as anti-cancer drugs is considered.
- PMID:
- 22281688
- [PubMed - as supplied by publisher]
Development of a human herpesvirus 6 virus species-specific immunoblotting assay.
Source
Department of Pediatrics, Fujita Health University School of Medicine.
Abstract
In order to assess the full spectrum of HHV-6A- and HHV-6B-associated diseases, we sought to develop a HHV-6 virus species-specific serological assay based on immunoblot analysis. The immunodominant proteins encoded by ORF U11, p100 for HHV-6A (strain U1102) and 101K for HHV-6B (strain Z29), were selected to generate virus species-specific antigens. Recombinant p100 and 101K were produced in a prokaryotic expression system. The expression of these proteins was confirmed using anti-His tag and 101K-specific monoclonal antibodies. HHV-6 virus species-specificantibodies were detected by immunoblotting in patient sera. Eighty seven serum samples obtained from various subjects were utilized to determine the reliability of the method for clinical use. Ten of 12 exanthem subitum convalescent sera reacted exclusively with 101K, while none of 12 acute sera reacted with either protein. Two of three sera collected from HHV-6A-infected patients reacted with p100 and 101K. Although all 5 acute and convalescent sera obtained from transplant recipients reacted exclusively with 101K, two of 6 convalescent sera obtained from patients with drug-induced hypersensitivity syndrome reacted with both p100 and 101K. Thirty-one of 38 sera obtained from healthy adults were positive for 101K antibody, while 4 reacted with both proteins. However, PCR analysis of PBMCs and saliva from these subjects did not detect HHV-6A DNA. In conclusion, this novel serological assay based on immunoblot analysis using recombinant HHV-6A p100 and HHV-6B 101K allowed to discriminate between HHV-6A- and HHV-6B-specific antibodies.
PAR-1-dependent and PAR-independent pro-inflammatory signalling in human lung fibroblasts exposed to thrombin.
Source
Centre for Respiratory Research, University College London, United Kingdom.
Abstract
Proteinase-activated receptors (PARs) are crucial in orchestrating cellular responses to coagulation proteinases, such as thrombin and FXa. Four PARs have been characterized and have been shown to be differentially expressed in mice and humans and between tissues. We have previously shown that in murine lung fibroblasts, PAR-1 is solely responsible for all cellular responses to thrombin and FXa. In contrast, we report here that in primary human lung fibroblasts, known PARs fail to account for all of the cellular responses to thrombin, in particular in the presence of high, but physiologically achievable concentrations of thrombin. We report that primary human lung fibroblasts secrete CCL2 in a PAR-1-dependent manner at low thrombin concentration (∼ 0.3 nM). At or above 10 nM thrombin, pharmacological antagonism (RWJ-58259) fails to block thrombin-induced CCL2 release; whereas PAR-1 cleavage-blocking monoclonal antibodies (ATAP2 and WEDE15) only partially inhibit thrombin-induced CCL2 secretion. In addition, activation of PAR-3, PAR-4 and transactivation of either PAR-2 or EGFR were ruled out as being responsible for thrombin-mediated CCL2 secretion at high yet standard concentrations of the proteinase. We further provide evidence that PAR-1-dependent and PAR-independent signaling involves the rapid phosphorylation of ERK which in turn is absolutely required for thrombin-induced CCL2 secretion at both low and standard concentration of the proteinase. Our findings suggest the existence of a PAR-independent signaling mechanism in human lung fibroblasts and have important implications for the design of therapeutic strategies aimed at blocking pro-inflammatory signaling responses associated with excessive thrombin generation. J. Cell. Physiol. © 2012 Wiley Periodicals, Inc.
Copyright © 2012 Wiley Periodicals, Inc.
Structural basis for broad detection of genogroup II noroviruses by amonoclonal antibody that binds to a site occluded in the viral particle.
Source
Department of Virology II, National Institute of Infectious Diseases, Tokyo, 208-0011, Japan.
Abstract
Human noroviruses are genetically and antigenically highly divergent. Monoclonal antibodies raised in mice against one kind of norovirus virus-like particle (VLP), however, were found to have broad recognition. In this study, we present the crystal structure of the antigen-binding fragment (Fab) for one of these broadly reactive monoclonal antibodies, 5B18, in complex with the capsid-protruding domain from a genogroup II genotype 10 (GII.10) norovirus at 3.3 Å resolution and also the cryo-electron microscopy structure of the GII.10 VLP at ∼10 Å resolution. The GII.10 VLP structure was more similar in overall architecture to the GV.1 murine norovirus virion than to the prototype GI.1 human norovirus VLP, with the GII.10 protruding domain raised ∼15 Å off the shell domain and rotated ∼40° relative to the GI.1 protruding domain. In the crystal structure, the 5B18 Fab bound to a highly conserved region of the protruding domain. Based on the VLP structure, this region is involved in interactions with other regions of the capsid and is partially buried in the virus particle. Despite the occluded nature of the recognized epitope in the VLP structure, ELISA binding suggested that the 5B18 antibody was able to capture intact VLPs. Together, the results provide evidence that the norovirus particle is capable of conformational flexibility, which may allow for antibody recognition of conserved surfaces that would otherwise be deeply buried on intact norovirus particles.
Pathogenic Anti-Desmoglein 3 mAbs Cloned from a Paraneoplastic Pemphigus Patient by Phage Display.
Source
1] Department of Dermatology, Keio University School of Medicine, Tokyo, Japan [2] Department of Dermatology, Cairo University School of Medicine, Cairo, Egypt.
Abstract
Paraneoplastic pemphigus (PNP) is an autoimmune blistering disease associated with lymphoproliferative neoplasms and characterized by antibodies against plakins and desmoglein 3 (Dsg3). Anti-Dsg3 antibodies have a primary role in blister formation in PNP. In this study, we used phage display to clone monoclonal anti-Dsg3 antibodies from a PNP patient to further characterize their pathogenicity. We isolated 20 unique Dsg3-reactive mAbs, which we classified into four groups according to the heavy-chain complementarity-determining region 3 (CDR3) region. Genetic analyses demonstrated that three antibody groups used the VH1-46 gene (18 clones) and one group used the VH1-02 gene (2 clones). The results of an in vitro keratinocyte dissociation assay and a human skin organ culture injection assay showed that three antibodies displayed pathogenic activity in blister formation with different potencies. Epitope mapping using domain-swapped Dsg3/Dsg2 showed that these pathogenic mAbs bound Ca(2+)-dependent conformational epitopes in the middle portion of the extracellular region of Dsg3 (EC2 and EC3 domains), in contrast to most previously characterized pathogenic pemphigus vulgaris antibodies, which bound to the EC1 domain of Dsg3. These mAbs reflect the unique polyclonal nature of anti-Dsg3 antibodies in PNP and represent an important tool for detailing the pathophysiological mechanisms of blister formation in PNP.Journal of Investigative Dermatology advance online publication, 26 January 2012; doi:10.1038/jid.2011.449.
Comparative analysis of carbohydrate-binding specificities of two anti-glycogen monoclonal antibodies using ELISA and surface plasmon resonance.
Source
Organization of Advanced Science and Technology, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657-8501, Japan.
Abstract
For immunological experiments on glycogens, anti-glycogen antibodies are indispensable to capture, detect, and visualize sugar molecules. An anti-glycogen monoclonal antibody (IV58B6) and newly constructed antibody (ESG1A9mAb) have a common immunoglobulin type (IgM) and binding ability to glycogens, but overall possess different binding features. Therefore, they may prove useful for the construction of an advanced system of quantitative ELISA based on their molecular structures. For this purpose, detailed information on the carbohydrate-specificities of ESG1A9mAb and IV58B6 is first required, but their fine specificities for various types of glycogens have not been elucidated. To overcome this problem, we performed interaction analysis by ELISA of ESG1A9mAb and IV58B6 toward 15 glucose polymers, that is, 5 enzymatically-synthesized glycogens (ESGs), 6 natural source glycogens (NSGs), 3 enzymatically digested glycogens (EDGs), and soluble starch. To provide a more detailed analysis, we determined the association constants (K(a)) of the two antibodies toward these glycogens by surface plasmon resonance. The results indicated that the carbohydrate-binding properties toward NSGs of ESG1A9mAb and IV58B6 were similar, but markedly differed for ESGs and EDGs. ESG1A9mAb showed significant affinity for all the ESGs and NSGs tested, whereas IV58B6 had only slight affinity for ESGs, although the affinities were increased when the ESGs were enzymatically digested. This information should be helpful for the design of both in vitro and in vivo immunological assays.
Copyright © 2012 Elsevier Ltd. All rights reserved.
Antibody therapy targeting ALS-linked misfolded protein.
Source
Molecular Neuroscience Research Center, Shiga University of Medical Science.
Abstract
Accumulating evidence indicates that the pathogenesis of Amyotrophic lateral sclerosis (ALS) is tightly linked to misfolding a key protein. Antibody therapy aims to eliminate or compete with the pathogenic proteins, through either passive or active immunization. We and others have generated several monoclonal antibodies (MAb)s which recognize only misfolded SOD1, but not the wild-type. Several MAbs are reported to delay the progression of mutant SOD1 Tg mice by the intraventricular application, which is mediated by different pathways. The determination of the pathogenic domain is crucial to acquire the effect of MAb therapy. Single chain of fragment of variance of IgG (scFv) is attracting emerging attention due to its broad application, in which intracellular proteins can be targeted by Intrabody or the modification of MAb with translocation signals.
Antibody therapy for Alzheimer's disease.
Source
Department of Diagnosis, Prevention and Treatment of Dementia, Graduate School of Medicine, Juntendo University.
Abstract
In order to avoid Abeta-induced autoimmune encephalitis, several monoclonal and polyclonal antibodies are in clinical trials. These are bapineuzumab, solanezumab, ponezumab, gantenerumab, BAN2401, gammaguard and octagam. Since each antibody has a different antigen epitope of Abeta, anti-amyloid activities are different. It is unknown which antibody is effective for Alzheimer disease, and we must wait for the result of clinical trials. Some patients who developed tissue amyloid plaque immuno-reactive (TAPIR) antibody showed slower decline after AN-1792 vaccination. We developed TAPIR-like monoclonal antibody, which was found to react with Abeta oligomers preferentially.
Single Domain Antibodies: A New Concept for Epidermal Growth Factor Receptor and EGFRvIII Targeting.
Source
Endocrine and Metabolism Research Center, Tehran University of Medical Sciences , Tehran, Islamic Republic of Iran .
Abstract
Epidermal growth factor receptor (EGFR) is one of the major molecular targets for cancer diagnosis and therapy. EGFR and EGFRvIII, mutated form of EGFR, have been identified as participating in pathogenesis of some forms of human cancers. Monoclonal antibodies (mAbs) targeting EGFR/EGFRvIII have been shown to suppress the signal transduction pathways controlling tumor cell growth, proliferation, and apoptosis. Until now, different types of mAbs or antibody fragments against EGFR family have been established. Some of these antibodies have been used clinically for treating various forms of human malignancies. More recently, a single domain antibody (sdAb) targeting this family of receptors has been introduced. The heavy chain antibodies (HCAbs) that made up variable regions of heavy chain, CH2, and CH3 domains are shown in camelids. SdAbs derived from camel HCAbs are the smallest known natural building parts for binding to antigen. They also possess a longer antigen recognizing region, which increases their capability for being more specific in target antigen enhancement. Camelid antibodies are highly valuable for their special characteristics, including heat resistance, small size, high solubility in an aqueous environment, and non-immunogenicity in a human environment. Due to these abilities, research on biotechnological production and treatment applications of recombinant smaller fragments of these only HCAbs is widely in progress. In this article, we will discuss the challenges and successes of different types of mAbs targeting EGFR/EGFRvIII in human cancer.
Antibody-induced enhancement of factor VIIa activity through distinct allosteric pathways.
Source
Aarhus University, Denmark;
Abstract
In the absence of its cofactor tissue factor (TF), coagulation factor VIIa (FVIIa) predominantly exists in a zymogen-like, catalytically incompetent state. Here we demonstrate that conformation-specific monoclonal antibodies (mAbs) can be used to characterize structural features determining the activity of FVIIa. We isolated two classes of mAbs, which both increased the catalytic efficiency of FVIIa more than 150-fold. The effects of the antibodies were retained with a FVIIa variant which has been shown to be inert to allosteric activation by the natural activator TF, suggesting that theantibodies and TF employ distinct mechanisms of activation. The antibodies could be classified into two groups based on their patterns of affinities for different conformations of FVIIa. Whereas one class of antibodies affected both the Km and kcat the other class mainly affected the Km. The antibody-induced activity enhancement could be traced to maturation of the substrate binding pockets and the oxyanion hole, evident by an increased affinity for p-aminobenzamidine, an increased rate of antithrombin inhibition, an increased rate of incorporation of diisopropylfluorophosphate, and an enhanced fraction of molecules with a buried N-terminus of the catalytic domain in the presence of antibodies. As demonstrated by site-directed mutagenesis, the two groups of antibodies appear to have overlapping, although clearly different, epitopes in the 170-loop. Our findings suggest that binding of ligands to specific residues in the 170-loop or its spatial vicinity may stabilize the S1 pocket and the oxyanion hole and they may have general implications for the molecular understanding of FVIIa regulatory mechanisms.
Select host cell proteins coelute with monoclonal antibodies in protein a chromatography.
Source
Dept. of Pharma Technical development, Genentech, Oceanside, CA 92056. bart.nogal@gmail.com.
Abstract
The most significant factor contributing to the presence of host cell protein (HCP) impurities in Protein A chromatography eluates is their association with the product monoclonal antibodies (mAbs) has been reported previously, and it has been suggested that more efficacious column washes may be developed by targeting the disruption of the mAbs-HCP interaction. However, characterization of this interaction is not straight forward as it is likely to involve multiple proteins and/or types of interaction. This work is an attempt to begin to understand the contribution of HCP subpopulations and/or mAb interaction propensity to the variability in HCP levels in the Protein A eluate. We performed a flowthrough (FT) recycling study with product respiking using two antibody molecules of apparently different HCP interaction propensities. In each case, the ELISA assay showed depletion of select subpopulations of HCP in Protein A eluates in subsequent column runs, while the feedstock HCP in the FTs remained unchanged from its native harvested cell culture fluid (HCCF) levels. In a separate study, the final FT from each molecule's recycling study was cross-spiked with various mAbs. In this case, Protein A eluate levels remained low for all but two molecules which were known as having high apparent HCP interaction propensity. The results of these studies suggest that mAbs may preferentially bind to select subsets of HCPs, and the degree of interaction and/or identity of the associated HCPs may vary depending on the mAb. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012.
Copyright © 2012 American Institute of Chemical Engineers (AIChE).
Developmental expression of Drosophila Wiskott-Aldrich Syndrome family proteins.
Source
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.
Abstract
BACKGROUND:
Wiskott-Aldrich Syndrome (WASP) family proteins participate in many cellular processes involving rearrangements of the actin cytoskeleton. To the date, four WASP subfamily members have been described in Drosophila: Wash, WASp, SCAR, and Whamy. Wash, WASp, and SCAR are essential during early Drosophila development where they function in orchestrating cytoplasmic events including membrane-cytoskeleton interactions. A mutant for Whamy has not yet been reported.
RESULTS:
We generated monoclonal antibodies that are specific to Drosophila Wash, WASp, SCAR, and Whamy, and use these to describe their spatial and temporal localization patterns. Consistent with the importance of WASP family proteins in flies, we find that Wash, WASp, SCAR, and Whamy are dynamically expressed throughout oogenesis and embryogenesis. For example, we find that Wash accumulates at the oocyte cortex. WASp is highly expressed in the PNS, while SCAR is the most abundantly expressed in the CNS. Whamy exhibits an asymmetric subcellular localization that overlaps with mitochondria and is highly expressed in muscle.
CONCLUSION:
All four WASP family members show specific expression patterns, some of which reflect their previously known roles and others revealing new potential functions. The monoclonal antibodies developed offer valuable new tools to investigate how WASP family proteins regulate actin cytoskeleton dynamics.
Copyright © 2012 Wiley Periodicals, Inc.
Risk management of biosimilars in oncology: each medicine is a work in progress.
Source
Hospital Pharmacy, Erasmus University Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands, a.vulto@erasmusmc.nl.
Abstract
Drug licensing and drug safety monitoring for standard chemical entities have been established and are routinely used. These have resulted in a solid foundation of knowledge from which confident therapeutic decisions can be made. For many chemical entities, this advanced level of experience is also present for the generic products. The expertise surrounding the development of biosimilar competitor versions is increasing and progress is encouraging. To address the re-engineering and comparability complexities of biosimilars, the European Union imposed a requirement that risk management plans be included in the medications' marketing applications. This paper summarizes and discusses the circumstances complicating the public's view of drug safety, historical incidents during the transition from innovative to competitor products, as well as retrospective assessments of the development and post-marketing experiences thus far with two biosimilars. Through assessing the market entries and post-marketing experiences of biosimilars used in oncology, the healthcare field can better prepare for the next wave of comparator-products: biosimilar monoclonal antibodies.
Newly Established Monoclonal Antibodies for Immunological Detection of H5N1 Influenza Virus.
Source
Department of Immunology, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.
Abstract
The H5N1 subtype of the highly pathogenic (HP) avian influenza virus has been recognized for its ability to cause serious pandemics among humans. In the present study, new monoclonal antibodies (mAbs) against viral proteins were established for the immunological detection of H5N1 influenza virus for research and diagnostic purposes. B-cell hybridomas were generated from mice that had been hyperimmunized with purified A/Vietnam/1194/2004 (NIBRG-14) virion that had been inactivated by UV-irradiation or formaldehyde. After screening over 4,000 hybridomas, eight H5N1-specific clones were selected. Six were specific for hemagglutinin (HA) and had in vitro neutralization activity. Of these, four were able to broadly detect all tested clades of the H5N1 strains. Five HA-specific mAbs detected denatured HA epitope(s) in Western blot analysis, and two detected HP influenza virus by immunofluorescence and immunohistochemistry. A highly sensitive antigen-capture sandwich ELISA system was established by combining mAbs with different specificities. In conclusion, these mAbs may be useful for rapid and specific diagnosis of H5N1 influenza. Therapeutically, they may have a role in antibody-based treatment of the disease.
Generation of anti-human DEC205/CD205 monoclonal antibodies that recognize epitopes conserved in different mammals.
Source
Laboratory of Cellular Physiology and Immunology and Chris Browne Center for Immunology and Immune Diseases, The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.
Abstract
DEC205/CD205 is a C-type multilectin receptor, expressed highly in dendritic cells (DCs). Previous efforts to generate anti-human DEC205 (anti-hDEC205) monoclonal antibodies (mAbs) from mice immunized with subdomain proteins of hDEC205 resulted in a few mAbs. Recently, we expressed and utilized a full-length extracellular domain protein of hDEC205 to successfully generate 5 strong anti-hDEC205 mAbs from mice. In this study, DEC205 knockout (KO) mice were immunized with this full-length extracellular domain protein of hDEC205. One of the 3 immunized DEC205 KO mice was chosen for the highest anti-hDEC205 titer by flow cytometric analysis of serum samples on CHO cells stably expressing hDEC205 (CHO/hDEC205 cells) and used for hybridoma fusion. From a single fusion, more than 400 anti-hDEC205 hybridomas were identified by flow cytometric screen with CHO/hDEC205 cells, and a total of 115 hybridomas secreting strong anti-hDEC205 mAb were saved and named HD1 through HD115. To characterize in detail, 10 HD mAbs were chosen for superior anti-hDEC205 reactivity and further subjected to cloning and purification. Interestingly, out of those 10 chosen anti-hDEC205 HD mAbs, 5 mAbs were also strongly reactive to mouse DEC205 while 8 mAbs were found to stain DEC205(+) DCs on monkey spleen sections. In addition, we also identified that HD83, one of the 10 chosen HD mAbs, stains DEC205(+) DCs in rat spleen and lymph node. Therefore, by immunizing DEC205 KO mice with a full-length extracellular domain protein of hDEC205, we generated a large number of strong anti-hDEC205 mAbs many of which are cross-species reactive and able to visualize DEC205(+) DCs in lymphoid tissues of other mammals.
Copyright © 2012. Published by Elsevier B.V.
Potential of T-regulatory cells to protect xenografts.
Source
aDivision of Clinical Immunology and Allergology, Department of Internal Medicine, University Hospital and Medical Faculty bGeneva Surgical Research Unit, Department of Surgery, University Hospital Geneva, Geneva cTransplantation Centre and Transplantation Immunopathology Laboratory, Department of Medicine, Centre Hospitalier Universitaire Vaudois (CHUV) and University of Lausanne, Lausanne, Switzerland.
Abstract
PURPOSE OF REVIEW:
Immunological barriers still preclude clinical xenotransplantation. The protective role of CD4CD25Foxp3 T-regulatory cells (Treg) in allotransplantation is well described and, therefore, could represent a promising therapeutical tool for xenotransplantation. This review addresses the latest findings on Treg in xenotransplantation research.
RECENT FINDINGS:
In vivo, costimulation blockade-based strategies including anti-CD154 monoclonal antibodies(mAbs) in combination with rapamycin or anti-LFA-1 mAb prolonged both concordant and discordant islets xenografts survival in a Treg-dependent manner. In vitro, IL-10 secretion was shown to be critical for the suppression of xenogeneic responses mediated by Treg. Moreover, transgenic expression of inducible costimulator-immunoglobulin or PD-L1 on porcine endothelial cells inhibited human T-cell proliferation in vitro and was associated with the induction of Treg and IL-10 secretion. CXCR3 mediated the recruitment of Treg to pig endothelium. Finally, the recruitment of human Treg was enhanced by the immobilization of human CCL17 on pig endothelium.
SUMMARY:
There is increasing evidence for the potential of CD4CD25Foxp3 Treg to protect xenografts. Induction of Treg in recipients and/or recruitment of human Treg to pig endothelium may represent novel strategies to prevent cell-mediated rejection in pig-to-human xenotransplantation.
Enzyme-Linked Immunosorbent Assay with Monoclonal and Single-Chain Variable Fragment Antibodies Selective to Coplanar Polychlorinated Biphenyls.
Abstract
Coplanar polychlorinated biphenyls (Co-PCBs) consisting of non-ortho and mono-ortho-chlorinated PCBs are dioxin-like compounds and caused contamination widely in the environment. In order to monitor Co-PCB residues, it was attempted to establish an enzyme-linked immunosorbent assay (ELISA) with monoclonal and recombinant antibodies selective to Co-PCBs. When 3,3',5,5'-tetrachlorobiphenoxy butyric acid (PCBH)-keyhole limpet hemocyanin conjugate was immunized into mice, two monoclonal antibodies Mab-0217 and Mab-4444 were obtained. 3,3',5,5'-Tetrachlorobiphenyl (PCB80) was determined with an IC50 value of 2.6 and 0.46 ng mL-1 in ELISA based on Mab-0217 and Mab-4444, respectively. Mab-4444 cross-reacted with Co-PCB congeners, except for PCB77 and PCB81. Mab-0217 reacted with PCB80 and cross-reacted with PCB111. A single-chain variable fragment (scFv) antibody derived from Mab-4444 was produced in recombinant E. coli cells. The scFv antibody showed nearly the same sensitivity toward PCBH as the parent monoclonal antibody in ELISA. These results clearly suggested that Mab-4444 and its scFv antibodies were suitable for monitoring of the representative congeners of Co-PCBs.
Anti-IL-2 Treatment Impairs the Expansion of T(reg) Cell Population during Acute Malaria and Enhances the Th1 Cell Response at the Chronic Disease.
Source
Departamento de Imunologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, São Paulo, Brasil.
Abstract
Plasmodium chabaudi infection induces a rapid and intense splenic CD4(+) T cell response that contributes to both disease pathogenesis and the control of acute parasitemia. The subsequent development of clinical immunity to disease occurs concomitantly with the persistence of low levels of chronic parasitemia. The suppressive activity of regulatory T (T(reg)) cells has been implicated in both development of clinical immunity and parasite persistence. To evaluate whether IL-2 is required to induce and to sustain the suppressive activity of T(reg) cells in malaria, we examined in detail the effects of anti-IL-2 treatment with JES6-1 monoclonal antibody (mAb) on the splenic CD4(+) T cell response during acute and chronic P. chabaudi AS infection in C57BL/6 mice. JES6-1 treatment on days 0, 2 and 4 of infection partially inhibits the expansion of the CD4(+)CD25(+)Foxp3(+) cell population during acute malaria. Despite the concomitant secretion of IL-2 and expression of high affinity IL-2 receptor by large CD4(+) T cells, JES6-1 treatment does not impair effector CD4(+) T cell activation and IFN-γ production. However, at the chronic phase of the disease, an enhancement of cellular and humoral responses occurs in JES6-1-treated mice, with increased production of TNF-α and parasite-specific IgG2a antibodies. Furthermore, JES6-1 mAb completely blocked the in vitro proliferation of CD4(+) T cells from non-treated chronic mice, while it further increased the response of CD4(+) T cells from JES6-1-treated chronic mice. We conclude that JES6-1 treatment impairs the expansion of T(reg) cell population during early P. chabaudi malaria and enhances the Th1 cell response in the late phase of the disease.
Fc gamma receptor 3a genotype predicts overall survival in follicular lymphoma patients treated on SWOG trials with combined monoclonalantibody plus chemotherapy but not chemotherapy alone.
Source
Arizona, USA;
Abstract
Background:Fc gamma receptor polymorphisms were linked to outcome in follicular lymphoma patients treated with single-agent rituximab, an anti-CD20 monoclonal antibody. In particular, 158F/F genotype of Fc gamma receptor 3A and 131R/R genotype of Fc gamma receptor 2A correlated with worse outcome compared to high-affinity 158V/V and 131H/H, respectively. We examined this association in the context of anti-CD20 monoclonal antibody combined with chemotherapy, as compared to chemotherapy alone, in follicular lymphoma patients treated on SWOG clinical trials.Design and Methods:Tissue from 142 SWOG patients treated with chemotherapy alone (protocol S8809, N=70) or combined chemotherapy and anti-CD20 monoclonal antibody (rituximab and Iodine I-131 tositumomab on protocols S9800 and S9911, N=30 and 42, respectively) was analyzed. DNA was extracted and assayed for Fc gamma receptor 3A V158F and 2A R131H polymorphisms using a TaqMan SNP assay. Stratified Cox regression was used to assess association with overall survival.Results:D. Persky et al. FRGR3A genotype predicts OS in FL treated with chemo + mAbFor Fc gamma receptor 3A there was an association with overall survival in the combination therapy trials but not in the chemotherapy-only trial. Having at least one Fc gamma receptor 3A V allele was associated with improved overall survival vs. F/F (HR = 0.33 (95% CI, 0.11, 0.96), p=0.042). For overall survival, there was evidence of a statistical interaction between the use of mAb and the number of V alleles (0, 1, or 2) (p=0.006), There was no such association for Fc gamma receptor 2A.Conclusions:Fc gamma receptor 3A polymorphism status may be predictive of survival in follicular lymphoma patients given treatments containing an anti-CD20 antibody but not treatment with chemotherapy alone. Thus, Fc gamma receptor 3A polymorphisms may be important to consider in designing new follicular lymphoma trials and new anti-CD20 monoclonal antibodies.(clinicaltrials.gov identifier: NCT00933127).
No comments:
Post a Comment