Review article: immunogenicity of anti-TNF biologics in IBD - the role of patient, product and prescriber factors.

BACKGROUND: Anti-drug antibodies (ADAs) to biologic therapies contribute to the loss of response and infusion reactions to anti-TNF drugs in patients with inflammatory bowel disease (IBD). The reasons behind this immunogenicity are complex, and have not been the focus of a dedicated review for prescribers. AIM: To provide an overview of the patient, product and prescriber factors, which have been associated with the immunogenicity of anti-TNF therapy, and draw conclusions for clinical practice. METHODS: Review of representative observational studies and clinical trials from the IBD and other literature, which report associations with ADA development, with a focus on infliximab and adalimumab. RESULTS: ADAs develop in 10-20% of patients receiving anti-TNF maintenance therapy, and these patients are three times more likely to lose response as ADA-negative patients. Patient genotype plays a role in ADA risk in a minority of patients, but age or disease type is not a major factor. Drug mishandling, such as agitation or freeze-thaw cycles, can induce protein aggregates, which are known to be immunogenic. Prescription of maintenance therapy with concomitant immunomodulators, and achieving suitable trough drug levels, reduces the risk of ADAs in patients with IBD. CONCLUSIONS: Patients and prescribers can take several steps to reduce the risk of development of anti-drug antibodies to anti-TNF antibodies. Further research is required to determine if immunogenic factors identified in other situations apply to use of anti-TNFs in IBD.

Knockdown of the glucocorticoid receptor alters functional integration of newborn neurons in the adult hippocampus and impairs fear-motivated behavior.

Glucocorticoids (GCs) secreted after stress reduce adult hippocampal neurogenesis, a process that has been implicated in cognitive aspects of psychopathology, amongst others. Yet, the exact role of the GC receptor (GR), a key mediator of GC action, in regulating adult neurogenesis is largely unknown. Here, we show that GR knockdown, selectively in newborn cells of the hippocampal neurogenic niche, accelerates their neuronal differentiation and migration. Strikingly, GR knockdown induced ectopic positioning of a subset of the new granule cells, altered their dendritic complexity and increased their number of mature dendritic spines and mossy fiber boutons. Consistent with the increase in synaptic contacts, cells with GR knockdown exhibit increased basal excitability parallel to impaired contextual freezing during fear conditioning. Together, our data demonstrate a key role for the GR in newborn hippocampal cells in mediating their synaptic connectivity and structural as well as functional integration into mature hippocampal circuits involved in fear memory consolidation.

Capillary electrophoresis-mass spectrometry using noncovalently coated capillaries for the analysis of biopharmaceuticals.

In this work, the usefulness of capillary electrophoresis-electrospray ionization time-of-flight-mass spectrometry for the analysis of biopharmaceuticals was studied. Noncovalently bound capillary coatings consisting of Polybrene-poly(vinyl sulfonic acid) or Polybrene-dextran sulfate-Polybrene were used to minimize protein and peptide adsorption, and achieve good separation efficiencies. The potential of the capillary electrophoresis-mass spectrometry (CE-MS) system to characterize degradation products was investigated by analyzing samples of the drugs, recombinant human growth hormone (rhGH) and oxytocin, which had been subjected to prolonged storage, heat exposure, and/or different pH values. Modifications could be assigned based on accurate masses as obtained with time-of-flight-mass spectrometry (TOF-MS) and migration times with respect to the parent compound. For heat-exposed rhGH, oxidations, sulfonate formation, and deamidations were observed. Oxytocin showed strong deamidation (up to 40%) upon heat exposure at low pH, whereas at medium and high pH, mainly dimer (>10%) and trisulfide formation (6-7%) occurred. Recombinant human interferon-ß-1a (rhIFN-ß) was used to evaluate the capability of the CE-MS method to assess glycan heterogeneity of pharmaceutical proteins. Analysis of this N-glycosylated protein revealed a cluster of resolved peaks which appeared to be caused by at least ten glycoforms differing merely in sialic acid and hexose N-acetylhexosamine composition. Based on the relative peak area (assuming an equimolar response per glycoform), a quantitative profile could be derived with the disialytated biantennary glycoform as most abundant (52%). Such a profile may be useful for in-process and quality control of rhIFN-ß batches. It is concluded that the separation power provided by combined capillary electrophoresis and TOF-MS allows discrimination of highly related protein species.

Mineralocorticoid receptors in control of emotional arousal and fear memory.

The stress hormone corticosterone acts via two receptor types in the brain: the mineralocorticoid (MR) and the glucocorticoid receptor (GR). Both receptors are involved in processing of stressful events. A disbalance of MR:GR functions is thought to promote stress-related disorders. Here we studied the effect of stress on emotional and cognitive behaviors in mice with forebrain-specific inactivation of the MR gene (MR(CaMKCre), 4 months old; and control littermates). MR(CaMKCre) mice responded to prior stress (5 min of restraint) with higher arousal and less locomotor activity in an exploration task. A fear conditioning paradigm allowed assessing in one experimental procedure both context- and cue-related fear. During conditioning, MR(CaMKCre) mice expressed more cue-related freezing. During memory test, contextual freezing remained potentiated, while control mice distinguished between cue (more freezing) and context episodes (less freezing) in the second memory test. At this time, plasma corticosterone levels of MR(CaMKCre) mice were 40% higher than in controls. We conclude that control of emotional arousal and adaptive behaviors is lost in the absence of forebrain MR, and thus, anxiety-related responses are and remain augmented. We propose that such a disbalance in MR:GR functions in MR(CaMKCre) mice provides the conditions for an animal model for anxiety-related disorders.

Corticosterone facilitates extinction of fear memory in BALB/c mice but strengthens cue related fear in C57BL/6 mice.

Corticosterone, the naturally occurring glucocorticoid of rodents is secreted in response to stressors and is known for its facilitating and detrimental effects on emotional learning and memory. The large variability in the action of corticosterone on processing of emotional memories is postulated to depend on genetic background and the spatio-temporal domain in which the hormone operates. To address this hypothesis, mice of two strains with distinct corticosterone secretory patterns and behavioural phenotype (BALB/c and C57BL/6J) were treated with corticosterone (250 microg/kg, i.p.), either 5 min before or directly after acquisition in a fear conditioning task. As the paradigm allowed assessing in one experimental procedure both context- and cue-related fear behaviour, we were able to detect generalization and specificity of fear. BALB/c showed generalized strong fear memory, while C57BL/6J mice discriminated between freezing during context- and cue episodes. Corticosterone had opposite effects on fear memory depending on the strain and time of injection. Corticosterone after acquisition did not affect C57BL/6J mice, but destabilized consolidation and facilitated extinction in BALB/c. Corticosterone 5 min before acquisition strengthened stress-associated signals: BALB/c no longer showed lower fear memory, while C57BL/6J mice displayed increased fear memory and impaired extinction in cue episodes. We propose that corticosterone-induced facilitation of fear memory in C57BL/6J mice can be used to study the development of fear memories, corticosterone administration in BALB/c mice presents a model to examine treatment. We conclude that genetic background and time of corticosterone action are modifiers of fear memory with interesting translational implications for anxiety-related diseases.

Strain specific fear behaviour and glucocorticoid response to aversive events: modelling PTSD in mice.

"Pavlovian" fear conditioning in rodents allows studying the formation and extinction of fear memories. Male C57BL/6J but not BALB/c mice showed differential fear memory performance expressed as freezing and scanning behaviour for context and cue. Glucocorticoid stress hormones modulate the processing of fear-related stimuli. The augmented corticosterone response of BALB/c mice to conditioning and testing, therefore, might have contributed to the strain-dependent formation of fear memories. We propose that modulation of extinction processes by glucocorticoids can be relevant in modelling anxiety disorders.