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In October 2013, the International Life Sciences Institute - Health and Environmental Sciences Institute Immunotoxicology Technical Committee (ILSI-HESI ITC) held a one-day workshop entitled, “Workshop on Cytokine Release: State-of-the-Science, Current Challenges and Future Directions”. The workshop brought together scientists from pharmaceutical, academic, health authority, and contract research organizations to discuss novel approaches and current challenges for the use of in vitro cytokine release assays (CRAs) for the identification of cytokine release syndrome (CRS) potential of novel monoclonal antibody (mAb) therapeutics. Topics presented encompassed a regulatory perspective on cytokine release and assessment, case studies regarding the translatability of preclinical cytokine data to the clinic, and the latest state of the science of CRAs, including comparisons between mAb therapeutics within one platform and across several assay platforms, a novel physiological assay platform, and assay optimization approaches such as determination of FcR expression profiles and use of statistical tests. The data and approaches presented confirmed that multiple CRA platforms are in use for identification of CRS potential and that the choice of a particular CRA platform is highly dependent on the availability of resources for individual laboratories (e.g. positive and negative controls, number of human blood donors), the assay through-put required, and the mechanism-of-action of the therapeutic candidate to be tested. Workshop participants agreed that more data on the predictive performance of CRA platforms is needed, and current efforts to compare in vitro assay results with clinical cytokine assessments were discussed. In summary, many laboratories continue to focus research efforts on the improvement of the translatability of current CRA platforms as well explore novel approaches which may lead to more accurate, and potentially patient-specific, CRS prediction in the future. [ABSTRACT FROM AUTHOR]

Abstract: Administration of biological therapeutic proteins can lead to unwanted immunogenicity in recipients of these products. The assessment and characterization of such immune reactions can be helpful to better understand their clinical relevance and how they relate to patient safety and therefore, have become an integral part of a product development program for biological therapeutics. Testing for anti-drug antibodies (ADA) to biological/biotechnology-derived therapeutic proteins generally follows a tiered approach. Samples are initially screened for binding antibodies; presumptive positives are then confirmed in a confirmatory assay; subsequently, confirmed-positive samples may be further characterized by titration and with a neutralizing antibody (NAb) assay. Regulatory guidances on immunogenicity state that assessing the neutralizing capacity of antibodies should preferably be done using functional bioassays, while recognizing that competitive ligand-binding (CLB) assays may be substituted when neutralizing bioassays are inadequate or not feasible. This manuscript describes case studies from four companies in which CLB assays and functional bioassays were compared for their ability to detect neutralizing ADA against a variety of biotechnology-derived therapeutic proteins. Our findings indicate that CLB assays are comparable to bioassays for the detection of NAbs, in some cases offering better detection sensitivity, lower variability, and less matrix interference. [ABSTRACT FROM AUTHOR]

Abstract: The administration of biological therapeutics may result in the development of anti-drug antibodies (ADAs) in treated subjects. In some cases, ADA responses may result in the loss of therapeutic efficacy due to the formation of neutralizing ADAs (NAbs). An important characteristic of anti-drug NAbs is their direct inhibitory effect on the pharmacological activity of the therapeutic. Neutralizing antibody responses are of particular concern for biologic products with an endogenous homolog whose activity can be potentially dampened or completely inhibited by the NAbs leading to an autoimmune-type deficiency syndrome. Therefore, it is important that ADAs are detected and characterized appropriately using sensitive and reliable methods. The design, development and optimization of cell-based assays used for detection of NAbs have been published previously by Gupta et al. 2007 . This paper provides recommendations on best practices for the validation of cell-based NAb assay and suggested validation parameters based on the experience of the authors. [Copyright &y& Elsevier]

This continuing education course was designed to provide an overview of the immunologic mechanisms involved in immunogenicity and hypersensitivity reactions following administration of biologics in nonclinical toxicity studies, the methods used to determine whether such reactions are occurring, and the associated clinical and anatomic pathology findings. Hypersensitivity reactions have classically been divided into type I, II, III, and IV reactions; type I and III reactions are those most often observed following administration of biologics. A variety of methods can be used to detect these reactions. Antemortem methods include hematology; detection of antidrug antibodies, circulating immune complexes and complement fragments, and immunoglobulin E in serum; tests for serum complement activity; and evaluation of complement receptor 1 on erythrocytes. Postmortem methods include routine light microscopy and electron microscopy, which can demonstrate typical findings associated with hypersensitivity reactions, and immunohistochemistry, which can detect the presence of immune complexes in tissues, including the detection of the test article. A final determination of whether findings are related to a hypersensitivity reaction in individual animals or across the entire study should rely on the overall weight of evidence, as findings indicative of these reactions are not necessarily consistent across all affected animals. [ABSTRACT FROM AUTHOR]

Abstract: Nerve growth factor (NGF) is an important mediator of pain and hyperalgesia and has become a target of novel analgesic therapeutics. Tanezumab is a humanized IgG2 antibody that binds NGF with high affinity and specificity. In a study to assess the toxicity and pharmacokinetic properties of tanezumab in adult, male and female, cynomolgus monkeys following weekly intravenous administration of 1, 10, or 30mg/kg for up to 26weeks (followed by an 8-week recovery period), tanezumab was well tolerated with no macroscopic or microscopic effects on those brain, spinal cord, nerve, or ganglia sections evaluated. One fifth of tanezumab-treated monkeys developed an antibody response to tanezumab that prevented maintenance of tanezumab exposure between dosing. In the antibody-negative animals, accumulation of tanezumab was observed; steady state was achieved approximately 8weeks after the first dose of study drug, and exposure to tanezumab was approximately dose proportional with no observed difference between male and female animals. One monkey died during the study; this monkey had findings suggestive of hypersensitivity reaction. The favorable toxicity and pharmacokinetic profile of tanezumab seen in this study supports its further evaluation for the treatment of pain in clinical practice. [Copyright &y& Elsevier]