Characterization of Molecular Determinants of the Conformational Stability of Macrophage Migration Inhibitory Factor: Leucine 46 Hydrophobic Pocket

Macrophage Migration Inhibitory Factor (MIF) is a key mediator of inflammatory responses and innate immunity and has been implicated in the pathogenesis of several inflammatory and autoimmune diseases. The oligomerization of MIF, more specifically trimer formation, is essential for its keto-enol tautomerase activity and probably mediates several of its interactions and biological activities, including its binding to its receptor CD74 and activation of certain signaling pathways. Therefore, understanding the molecular factors governing the oligomerization of MIF and the role of quaternary structure in modulating its structural stability and multifunctional properties is crucial for understanding the function of MIF in health and disease. Herein, we describe highly conserved intersubunit interactions involving the hydrophobic packing of the side chain of Leu46 onto the β-strand β3 of one monomer within a hydrophobic pocket from the adjacent monomer constituted by residues Arg11, Val14, Phe18, Leu19, Val39, His40, Val41, Val42, and Pro43. To elucidate the structural significance of these intersubunit interactions and their relative contribution to MIF’s trimerization, structural stability and catalytic activity, we generated three point mutations where Leu46 was replaced by glycine (L46G), alanine (L46A) and phenylalanine (L46F), and their structural properties, stability, oligomerization state, and catalytic activity were characterized using a battery of biophysical methods and X-ray crystallography. Our findings provide new insights into the role of the Leu46 hydrophobic pocket in stabilizing the conformational state of MIF in solution. Disrupting the Leu46 hydrophobic interaction perturbs the secondary and tertiary structure of the protein but has no effect on its oligomerization state

Circulating CD56^bright NK cells inversely correlate with survival of melanoma patients.

The roles of NK cells in human melanoma remain only partially understood. We characterized NK cells from peripheral blood ex vivo by flow cytometry obtained from late stage (III/IV) melanoma patients. Interestingly, we found that the abundance of CD56 <sup>bright</sup> NK cells negatively correlate with overall patient survival, together with distant metastases, in a multivariate cox regression analysis. The patients' CD56 <sup>bright</sup> NK cells showed upregulation of CD11a, CD38 and CD95 as compared to healthy controls, pointing to an activated phenotype as well as a possible immune regulatory role in melanoma patients. After stimulation in vitro, CD56 <sup>bright</sup> NK cells produced less TNFα and GMCSF in patients than controls. Furthermore, IFNγ production by the CD56 <sup>bright</sup> NK cells correlated inversely with overall survival. Our results highlight that abundance and function of CD56 <sup>bright</sup> NK cells are associated with melanoma patient survival, emphasizing the potential of NK cell subsets for biomarker discovery and future therapeutic targeting

Macrophage Migration Inhibitory Factor Is Enhanced in Acute Coronary Syndromes and Is Associated with the Inflammatory Response

Chronic inflammation promotes atherosclerosis in cardiovascular disease and is a major prognostic factor for patients undergoing percutaneous coronary intervention (PCI). Macrophage migration inhibitory factor (MIF) is involved in the progress of atherosclerosis and plaque destabilization and plays a pivotal role in the development of acute coronary syndromes (ACS). Little is known to date about the clinical impact of MIF in patients with symptomatic coronary artery disease (CAD).In a pilot study, 286 patients with symptomatic CAD (n = 119 ACS, n = 167 stable CAD) undergoing PCI were consecutively evaluated. 25 healthy volunteers served as control. Expression of MIF was consecutively measured in patients at the time of PCI. Baseline levels of interleukin 6 (IL-6), “regulated upon activation, normal T-cell expressed, and secreted” (RANTES) and monocyte chemoattractant protein-1 (MCP-1) were measured by Bio-Plex Cytokine assay. C-reactive protein (CRP) was determined by Immunoassay. Patients with ACS showed higher plasma levels of MIF compared to patients with stable CAD and control subjects (median 2.85 ng/mL, interquartile range (IQR) 3.52 versus median 1.22 ng/mL, IQR 2.99, versus median 0.1, IQR 0.09, p<0.001). Increased MIF levels were associated with CRP and IL-6 levels and correlated with troponin I (TnI) release (spearman rank coefficient: 0.31, p<0.001). Patients with ACS due to plaque rupture showed significantly higher plasma levels of MIF than patients with flow limiting stenotic lesions (p = 0.002).To our knowledge this is the first study, demonstrating enhanced expression of MIF in ACS. It is associated with established inflammatory markers, correlates with the extent of cardiac necrosis marker release after PCI and is significantly increased in ACS patients with “culprit” lesions. Further attempts should be undertaken to characterize the role of MIF for risk assessment in the setting of ACS

Small-molecule inhibitors of macrophage migration inhibitory factor(MIF) as an emerging class of therapeutics for immune disorders

Macrophage migration inhibitory factor (MIF) is an important cytokine for which an increasing number of functions is being described in the pathogenesis of inflammation and cancer. Nevertheless, the availability of potent and druglike MIF inhibitors that are well-characterized in relevant disease models remains limited. Development of highly potent and selective small-molecule MIF inhibitors and validation of their use in relevant disease models will advance drug discovery. In this review, we provide an overview of recent advances in the identification of MIF as a pharmacological target in the pathogenesis of inflammatory diseases and cancer. We also give an overview of the current developments in the discovery and design of small-molecule MIF inhibitors and define future aims in this fiel

Macrophage migration inhibitory factor is critically involved in basal and fluoxetine-stimulated adult hippocampal cell proliferation and in anxiety, depression, and memory-related behaviors.

Intensive research is devoted to unravel the neurobiological mechanisms mediating adult hippocampal neurogenesis, its regulation by antidepressants, and its behavioral consequences. Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that is expressed in the CNS, where its function is unknown. Here, we show, for the first time, the relevance of MIF expression for adult hippocampal neurogenesis. We identify MIF expression in neurogenic cells (in stem cells, cells undergoing proliferation, and in newly proliferated cells undergoing maturation) in the subgranular zone of the rodent dentate gyrus. A causal function for MIF in cell proliferation was shown using genetic (MIF gene deletion) and pharmacological (treatment with the MIF antagonist Iso-1) approaches. Behaviorally, genetic deletion of MIF resulted in increased anxiety- and depression-like behaviors, as well as of impaired hippocampus-dependent memory. Together, our studies provide evidence supporting a pivotal function for MIF in both basal and antidepressant-stimulated adult hippocampal cell proliferation. Moreover, loss of MIF results in a behavioral phenotype that, to a large extent, corresponds with alterations predicted to arise from reduced hippocampal neurogenesis. These findings underscore MIF as a potentially relevant molecular target for the development of treatments linked to deficits in neurogenesis, as well as to problems related to anxiety, depression, and cognition