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Influence of Glycosaminoglycans on the Transmetallation and Transchelation Kinetics of Gadolinium from GBCAs in the Presence of ZnCl₂

Patrick Werner¹, Patrick Schuenke², Matthias Taupitz³, and Leif Schröder¹
¹Deutsches Krebsforschungszentrum, Heidelberg, Germany, ²Physikalisch-Technische Bundesanstalt, Berlin, Germany, ³Charite, Berlin, Germany

Synopsis

Relaxation rates in aqueous solutions containing 30-100 µM heparin as well as 150 µM GBCA were measured as a function of time. Different ZnCl₂ stimuli with concentrations between 0.125-4 mM were used as competing ions that initiate a transmetallation and a transchelation process of the Gd³⁺ ion from GBCAs to glycosaminoglycans. The time resolved relaxometry measurements indicate that glycosaminoglycans play a concentration-dependent double role as competing chelator structures. They foster the thermodynamic instability of intact GBCA by sequestering Gd³⁺ from the contrast agent but simultaneously interact with competing ions and thus cause a reduced kinetic instability.

Introduction

Clinically used Gadolinium-based contrast agents (GBCA) require a high stability of the Gd³⁺ complex even in the presence of competing ions and/or chelators. However, the accumulation of Gd³⁺ ions in the human body^1-3 was repeatedly observed on T₁-weighted images after the administration of GBCA^4-6. Insights about the exact mechanism of Gd³⁺ deposition in various tissues remain, however, elusive. Sugar structures like glycosaminoglycans (GAGs) are potential candidates to enlighten this missing piece². This study aims to elucidate the influence of different heparin concentrations on the kinetic stability of GBCAs in the presence of different ZnCl₂ concentrations.

Methods

We studied solutions with different concentrations of heparin (0 µM-100 µM) and ZnCl₂ (0.125-4 mM) in combination with five linear GBCAs (150 µM). All time resolved MR measurements were performed on a vertical 9.4 T MRI system. R₁ measurements at 25 °C were performed using a dephasing recovery sequence. All R₁ values reflect ROI-averaged values from corresponding R₁ parameter maps.

Results

A release of Gd³⁺ from GBCAs that is triggered by Zn²⁺ ions can be observed via an increase in the relaxation rate. In this process, part of the low relaxivity contribution of the GBCAs is replaced with the higher relaxivity contribution of Gd³⁺ in water or from the Gd³⁺ GAG complexes⁷. The determined time constants for the transmetallation process vary significantly between the different GBCAs and for the investigated ZnCl₂ concentrations (Tab.1). The final R₁ values after the transmetallation (plateau values of the curves) increase with increasing ZnCl₂ concentrations. A rather special behavior was observed for Primovist^® where the data does not reach a stable plateau during the observation time window. The time constants for the transchelation process in the aqueous heparin solutions differ significantly from the time constants for the transmetallation process in water (Tab.1). While the time constants for the transmetallation process were in the order of 2 minutes and slightly increased with increasing ZnCl₂ concentrations, the time constants for the transchelation are in the order of hours to weeks and tremendously decrease with increasing ZnCl₂ concentrations.

Discussion

Our investigations by time resolved MR relaxometry could show that the Zn/GAG ratio is critical to regain faster exchange kinetics during the transchelation and transmetallation process of Gd³⁺ from GBCAs. More Zn²⁺ ions potentially reduce the availability of binding sites in heparin. It was observed that the overall transchelation process is significantly more efficient for a high Zn/GAG ratio (Tab.1). The explanation could be that the larger ZnCl₂ concentration causes on average more Zn²⁺ to be outside the GAG and thus encounter GBCA complexes with a subsequent quick transmetallation step. Most importantly, contrary to the relatively fast displacement of Gd³⁺ by Zn²⁺ in the absence of competing chelators, the overall transchelation of Gd³⁺ to heparin needs up to multiple days to achieve a new chemical equilibrium (Tab.1 and Fig.2). This reflects the complex situation when Gd³⁺ ions and other cations compete for both the binding in the parent GBCA chelator and the binding sites in the GAG. It could be concluded that for small ZnCl₂ stimuli, the transmetallation is the limiting step and not the availability of binding sites for the subsequent chelation of dissociated Gd³⁺ to GAGs. However, from the transmetallation experiments in aqueous solutions, it was shown that the addition of ZnCl₂ causes the release of Gd³⁺ ions within a few minutes (Tab.1 and Fig.1). Hence, the stability of the GBCA complex per se cannot be the limiting factor either. It is thus further concluded that the Zn²⁺ ions are sufficiently withheld by their own binding to heparin from initiating the transmetallation of the GBCA. For strong ZnCl₂ stimuli, the time constant drops significantly. Due to the above-mentioned fact that Gd³⁺ outperforms Zn²⁺ in terms of binding to the GAG, the released Gd³⁺ions are apparently not limited in finding a binding site and forming the macromolecular Gd-GAG complexes.

Conclusion

We were able to demonstrate that GAGs as endogenous sugars play an important double role as competing chelators. Firstly, they foster the thermodynamic instability of intact GBCA in vitro by sequestering Gd³⁺ from the disfavored ZnL + Gd³⁺ intermediate. At the same time, the GAG’s interaction with competing ions can suppress the initial attack and reduce kinetic instability significantly. These findings add an important aspect to the ongoing discussion of gadolinium depositions in the human body after the application of GBCAs.

Acknowledgements

This research was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation)—Grant No. 289347353(GRK 2260) and Koselleck Grant No. 316693477 (SCHR 995/5–1). Support by the Dieter Morszeck Stiftung is also gratefully acknowledged.

References

1) Laurent, S., Elst, L. V, Copoix, F. & Muller, R. N. Stability of MRI paramagnetic contrast media: a proton relaxometric protocol for transmetallation assessment. Invest. Radiol. 36, 115–22 (2001).

2) Taupitz, M. et al. Gadolinium-containing magnetic resonance contrast media: investigation on the possible transchelation of Gd3+ to the glycosaminoglycan heparin. Contrast Media Mol. Imaging 8, 108–16 (2013).

3) Gianolio, E., Gregorio, E. Di & Aime, S. Chemical Insights into the Issues of Gd Retention in the Brain and Other Tissues Upon the Administration of Gd-Containing MRI Contrast Agents. Eur. J. Inorg. Chem. 2019, 137–151 (2019).

4) Kanda, T., Ishii, K., Kawaguchi, H., Kitajima, K. & Takenaka, D. High signal intensity in the dentate nucleus and globus pallidus on unenhanced T1-weighted MR images: relationship with increasing cumulative dose of a gadolinium-based contrast material. Radiology 270, 834–41 (2014).

5) Kanda, T. et al. High Signal Intensity in Dentate Nucleus on Unenhanced T1-weighted MR Images: Association with Linear versus Macrocyclic Gadolinium Chelate Administration. Radiology 275, 803–9 (2015).

6) Radbruch, A. et al. Gadolinium retention in the dentate nucleus and globus pallidus is dependent on the class of contrast agent. Radiology 275, 783–91 (2015).

7) Werner, P., Taupitz, M., Schröder, L., & Schuenke, P. An NMR relaxometry approach for quantitative investigation of the transchelation of gadolinium ions from GBCAs to a competing macromolecular chelator. Scientific Reports, 11(1), 1-13 (2021).

Figures

Figure 1: Time-resolved R₁ measurements of 150 µM A Optimark^®, B Omniscan^®, C Magnevist^®, D MultiHance^® and E Primovist^® in water after the addition of different ZnCl₂ concentrations between 125 µM and 4 mM. The black dashed lines represents the theoretical value of 150 µM GBCA in water. Only the values of Magnevist^® and MultiHance^® could be recorded for all ZnCl₂ concentrations. Omniscan^® and Optimark^® show a significantly faster dynamic than the other contrast media and Primovist^®was the contrast agent with the highest kinetic stability under these experimental circumstances.

Figure 2: Comparison of the transchelation kinetics of 150 µM Optimark^® A-C, Omniscan D-F, Magnevist^® G-I, MultiHance^® J-L and Primovist^® M-O in 100 µM, 65 µM and 30 µM heparin solution, respectively. The black dashed lines represent the value of 150 µM GBCA in heparin solution. All linear dianionic GBCAs show a significantly slower transchelation dynamic. In general, it can be seen that with decreasing heparin content, the transchelation dynamics of all GBCAs become faster and more Gd³⁺ ions are dissociated in shorter time.

Table 1: Determined time constants for the dynamic transmetallation (τ in H₂O) and dynamic transchelation (τ in 30 µM, 65 µM and 100 µM heparin) experiments.

Glycosaminoglycans as endogenous sugar structures foster the thermodynamic instability of intact GBCAs and at the same time, interact with competing ions which suppress the initial attack and reduce kinetic instability of GBCAs significantly.

Proc. Intl. Soc. Mag. Reson. Med. 30 (2022)

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DOI: https://doi.org/10.58530/2022/1442