LETTER AND REPLY
`
`Port J Nephrol Hypert 2012; 26(4): 000-000
`Advance Access publication 27 September 2012
`
`Assessment of dextran antigenicity of intravenous iron products by an immunodiffusion assay
`Susann Neiser, Maria Wilhelm, Katrin Schwarz, Felix Funk, Peter Geisser, Susanna Burckhardt
`Vifor (International) Inc. St. Gallen, Switzerland
`
`Port J Nephrol Hypert 2011; 25(3): 219-224
`Advance Access publication 13 September 2011
`
`LETTER (cid:132)
`
`Johannes Ring and Rudi Valenta
`
`Klinik und Poliklinik für Dermatologie und Allergologie am Biederstein der TU München
`München, Germany.
`
`Received for publication:
`Accepted:
`
`04/06/2012
`11/06/2012
`
`Sir,
`
`We have read the article by Susann Neiser et al. in a previous issue of this journal 2011;25(3):219-224
`with the title: “Assessment of dextran antigenicity of intravenous iron products by an immunodiffusion
`assay” and feel motivated to comment on it critically since we feel that the methodology and conclusions
`contain some errors which fundamentally challenge the interpretation of the findings.
`
`The clinical relevance: The assessment of the antigenicity of an injectable molecule for the human
`organism requires human patient sera due to the inherent variability of antibodies from person to
`person, and the inherent variation of bioactivity of antibodies within a single individual. Antibodies
`produced experimentally in animals give little information with regard to clinical effects of the antigenic
`structure.
`
`The authors postulate in their conclusion that “the reported immunoassay represents a possible
`approach for the evaluation of the risk of DIAR (dextran induced anaphylactic reactions)”. Dextran
`reactive antibodies have been intensively investigated in the past due to rare allergic reactions to
`large doses of high molecular weight dextrans used as plasma expanders1-5. These studies showed
`that the vast majority of normal volunteers have detectable levels of anti-dextran antibodies in their
`blood, yet the risk of clinically significant anaphylactoid reactions is well below 0.1%. Although some
`degree of correlation was found between the antibody titre (i.e. concentration) and the risk of ana-
`phylaxis among patients exposed to dextran infusions, it was however concluded that “dextran reactive
`antibodies per se have no pathogenic importance, since the great majority of volunteers with dextran
`reactive antibodies tolerate high molecular weight dextran infusion6. The true biological mechanism
`of dextran related anaphylaxis is explained as immunocomplex anaphylaxis with some still unknown
`aspects.
`
`The antibodies used: The authors postulate that antibodies prepared under highly artificial conditions in
`experimental animals can be used to predict the risk of clinical allergic reactions in patients. It is well known
`
` 1
`
`Pharmacosmos, Exh. 1045, p. 1
`
`

`
`Johannes Ring, Rudi Valenta
`
`from any textbook of immunology that experimental immunisation using standard adjuvants is the standard
`procedure of today, to provoke antibody reactions that would not otherwise occur without such artificial
`procedure7. To further underline the inappropriateness of the present article, the methodology section does
`not contain any information on the antibody specificity or immunisation procedure used, which also raises
`questions, since all data in the article is based on this one experimental antibody.
`
`The methodology used: The authors apply the radial immunodiffusion technology for the investiga-
`tion of immune reactions between IV iron compounds and the experimental antibody. This technology
`from the 1970s is rarely used today due to its low sensitivity. Analytical techniques for precise char-
`acterisation of anti-drug antibodies (ADAs) have developed considerably over the last few decades due
`to the need to measure neutralising antibodies to widely used genetically engineered protein drugs
`such as insulin or the TNF-α inhibitors8. The frequent occurrence of anti-TNF-α inhibitor antibodies in
`patients does not generally lead to anaphylactic events, but may result in rapid removal of the drug
`from the circulation resulting in reduced half-life, reduced bioactivity and reduced clinical efficacy9.
`Today’s standards for analysis of ADAs are highly sensitive radio-immunoassays (RIAs) or highly sensi-
`tive enzyme linked immune-sorbent assays (ELISAs) which also enable analytical detection of subtype
`specific antibodies.
`
`It is regrettable that the authors have chosen an almost obsolete methodology and that the experi-
`mental setup is lacking proper controls which makes it difficult or impossible to judge the significance
`of the very faint precipitation patterns observed. It is thus not possible from the hardly visible precipi-
`tations to conclude whether they represent artefacts or true reactions. Under normal circumstances it
`is also required that precipitation rings in immunodiffusion analysis are clearly separated from the
`application well in order to be judged a positive reaction. This requirement is unfortunately not fulfilled
`in the present publication.
`
`Bioactivity of antibodies: It is well known from any textbook of basic immunology7 that the mere
`presence of antibodies does not imply a particular pathogenic or protective immune reaction or
`biological activity. The bioactivity of any antibody molecule is variable and linked to the constant
`part (Fc-fragment) of the antibody molecule, which possesses the receptor sites for initiation of
`bioactivity. This bioactivity may vary from none – if the antibody is responsible for conferring toler-
`ability – to induction of inflammation or anaphylaxis if the antibody molecule has the ability to bind
`to specific receptors on cell surfaces like high affinity FER on mast cells or basophils or to form antibody-
`antigen aggregates of sufficient size to activate the complement system or trigger the various other
`cascades.
`
`The team of authors: All the authors are affiliated to a company (Vifor) producing products competing
`with dextran-containing products. This may represent a conflict of interest in interpretation of test
`results.
`
`Conflict of interest statement.
`The authors of the letter have no conflict of interests. They are not financially connected with any company possibly involved with
`the content of either the article in question or this Letter to the Editor.
`
`2 Port J Nephrol Hypert 2012; 26(4): 000-000
`
`Pharmacosmos, Exh. 1045, p. 2
`
`

`
`Letter
`
`References
`
` 1. Ring J, Messmer K. Incidence and severity of anaphylactoid reactions to colloid volume
`substitutes. Lancet 1977; i466
`
` 2. Seemann C, Hedin H, Richter W, Ring J, Stippig S, Messmer K. Hapten-inhibition of
`dextran-induced anaphylactoid reaction. Eur Surg Res 1978;10:295
`
` 3. Ring J. Anaphylactoid reactions to intravenous solutions used for volume substitution.
`Clin Rev Allergy 1991;187:397-414
`
` 4. Kraft D, Hedin H, Richter W, Scheiner O, Rumpold H, Devey M.E. Immunoglobulin
`Class and Subclass Distribution of Dextran-Reactive Antibodies in Human Reactors
`and Non Reactors to Clinical Dextran. Allergy 1982;37: 481-489
`
` 5. Ljungström KG. Safety of dextran in relation to other colloids - 10 years experience
`with hapten inhibition. Infusionsther Tansfusionsmed 1993;20:206-210
`
` 6. Richter AW, Hedin HI. Dextran hypersensitivity. Immunology Today 1982;3:132-8
`
` 7. Goldsby, RA, Kindt, TJ, Osborne BA., Kuby, J. Antigens. In: Goldsby R.A, Kindt T.J,
`Kuby J, Osbourne BA, eds. “Immunology”, 5th ed. New York: W.H.Freeman & Co,
`2003; 57-74
`
` 8. Svenson M, Geborek P, Saxne T, Bendtzen K. Monitoring patients treated with anti-
`TNF-alpha biopharmaceuticals: assessing serum infliximab and anti-infliximab antibod-
`ies. Rheumatology (Oxford) 2007;46:1828-34
`
` 9. Bendtzen K, Ainsworth M, Steenholdt C, Thomsen OØ, Brynskov J. Individual medicine
`in inflammatory bowel disease: monitoring bioavailability, pharmacokinetics and
`immunogenicity of anti-tumour necrosis factor-alpha antibodies. Scand J Gastroen-
`terol 2009;44:774-81
`
`Correspondence to:
`Prof. Johannes Ring
`Klinik und Poliklinik für Dermatologie und Allergologie
`am Biederstein der TU München
`Biedersteiner Str. 29
`80802 München, Germany
`EMail: johannes.ring@lrz.tum.de
`
`REPLY
`
`Port J Nephrol Hypert 2012; 26(4): 000-000 3
`
`Pharmacosmos, Exh. 1045, p. 3