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Updated: May 19, 2026
In Vitro Assay to Study Tumor-macrophage Interaction
Published on: August 1, 2019
Structural interactions dictate the kinetics of macrophage migration inhibitory factor inhibition by different
Gregg V Crichlow1, Chengpeng Fan, Camille Keeler
1Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06510, USA.
Abstract:
Regulation of cellular processes by dietary nutrients is known to affect the likelihood of cancer development. One class of cancer-preventive nutrients, isothiocyanates (ITCs), derived from the consumption of cruciferous vegetables, is known to have various effects on cellular biochemistry. One target of ITCs is macrophage migration inhibitory factor (MIF), a widely expressed protein with known inflammatory, pro-tumorigenic, pro-angiogenic, and anti-apoptotic properties. MIF is covalently inhibited by a variety of ITCs, which in part may explain how they exert their cancer-preventive effects. We report the crystallographic structures of human MIF bound to phenethylisothiocyanate and to l-sulforaphane (dietary isothiocyanates derived from watercress and broccoli, respectively) and correlate structural features of these two isothiocyanates with their second-order rate constants for MIF inactivation. We also characterize changes in the MIF structure using nuclear magnetic resonance heteronuclear single-quantum coherence spectra of these complexes and observe many changes at the subunit interface. While a number of chemical shifts do not change, many of those that change do not have features similar in magnitude or direction for the two isothiocyanates. The difference in the binding modes of these two ITCs provides a means of using structure-activity relationships to reveal insights into MIF biological interactions. The results of this study provide a framework for the development of therapeutics that target MIF.
Insights
Dietary nutrients like isothiocyanates (ITCs) can prevent cancer by inhibiting macrophage migration inhibitory factor (MIF). Structural analysis reveals how ITCs bind to MIF, offering insights for developing new cancer therapeutics.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- Dietary nutrients, particularly isothiocyanates (ITCs) from cruciferous vegetables, play a role in cancer prevention.
- Macrophage migration inhibitory factor (MIF) is a protein implicated in inflammation, tumor growth, angiogenesis, and apoptosis resistance.
- ITCs exert cancer-preventive effects, partly through covalent inhibition of MIF.
Purpose of the Study:
- To elucidate the structural basis of MIF inhibition by dietary ITCs.
- To correlate structural features of ITCs with their efficacy in MIF inactivation.
- To understand the molecular interactions between ITCs and MIF for therapeutic development.
Main Methods:
- Crystallographic structure determination of human MIF complexed with phenethylisothiocyanate and l-sulforaphane.
- Kinetic analysis to determine second-order rate constants for MIF inactivation by ITCs.
- Nuclear Magnetic Resonance (NMR) spectroscopy (heteronuclear single-quantum coherence) to characterize structural changes in MIF upon ITC binding.
Main Results:
- Determined crystal structures of human MIF bound to phenethylisothiocyanate and l-sulforaphane.
- Correlated structural characteristics of these ITCs with their MIF inactivation rates.
- NMR analysis revealed significant structural changes at the MIF subunit interface, with distinct patterns for each ITC.
- Observed differences in binding modes and structural perturbations between the two ITCs.
Conclusions:
- The distinct binding modes of phenethylisothiocyanate and l-sulforaphane to MIF provide structure-activity relationship insights.
- Understanding these interactions is crucial for developing targeted therapeutics against MIF.
- This study lays the groundwork for novel therapeutic strategies aimed at inhibiting MIF's pro-tumorigenic functions.
