Synopsis

BAY 1436032 is a small-molecule inhibitor of the R132X-mutant form of IDH1 with a trifluoromethoxy-group that is currently clinically investigated in IDH1-mutant solid tumors. Non-invasive monitoring of intracranial drug delivery of BAY 1436032 in an animal model using ¹⁹F MRS is technically challenging but feasible.

Introduction

With 70–80% of astrocytomas and oligodendrogliomas harboring mutations in IDH1 genes, the mutant form of the enzyme is an attractive therapy target.^{1 2} Accumulating intracellular levels of (D-)2-Hydroxyglutarate (2-HG) in mutated cells presumably lead to a block in differentiation of non-transformed cells.³ BAY 1436032 is a small-molecule inhibitor of the R132X-mutant form of IDH1, inhibiting the mutation mediated generation of 2-HG. Since the inhibitor has a trifluoromethoxy-group (Fig.1) and the endogenous ¹⁹F MRI signal from the body is considered to be negligible, intracranial drug delivery might be monitored using ¹⁹F NMR/MR-Spectroscopy.

Results

We were able to record five fluor signal peaks in patient serum samples using NMR spectroscopy, with one specifically identified as albumin after titration with HSA (Fig.3). Spectra were externally referenced to TFA at -76.55 ppm with the ¹⁹F signal of the inhibitor identified at approximately -59 ppm (free compound). 3/10 mice in the GL261 group had to be sacrificed prior to treatment due to acute and severe clinical symptoms and 2/10 in the LNT-229 IDH1R132H due to suspected meningeosis. Anatomical scans on day 26 for LNT-229 revealed tumor take in 10/10 animals, on day 33 for LNT-229 IDH1R132H in 8/10 animals and on day 23 for GL261 in 7/7 (remaining) animals. Four mice in the GL261 group and one non-tumor bearing mouse died during single pulse sequence. ¹⁹F signal could be identified in most mice at approximately -59 ppm to -60 ppm. Signal shape revealed one rather sharp w/o multiple broad peaks. Excluding animals that died during the single pulse sequence, ¹⁹F signal amplitudes for largely protein bound inhibitor showed a low positive correlation (Pearson‘s r = 0.32) with total tissue concentration determined by MS (Fig. 4). Total concentration in brain was not dependent on tumor model. Phantom experiments recording the water signal (pulse FID) were conducted to compare water signal intensity (110 [M]) to fluor signal intensity (10 [M]). Based on this calibration, approx. 1/100^th of BAY 1436032 concentration in in vivo experiments can be detected by ¹⁹F MRS. Evaluation of 1H MRS showed a significant decrease in 2-HG/Cr ratios (p = 0.04; Fig. 5) from baseline to post-treatment follow-up scans in the LNT-229 IDH1R132H tumor model. SNR at 3T in patients was not sufficient for ¹⁹F signal detection.

Discussion

Despite a low signal intensity of a substance with only three fluor atoms which is mainly protein bound⁴, ¹⁹F MRS signal of BAY 1436032 can be identified intracranial in an animal model using a ¹⁹F single pulse sequence at a 300 MHz small animal scanner. We were not able to detect differences in concentrations of mice bearing invasive tumor models with a glioblastoma phenotype and a relevant BBB integrity loss (GL261) and with a less severe breakdown in slower growing models (LNT-229). This suggests that BBB breakdown might not affect the activity of a systemic therapy with BAY 1436032. In literature there is still an ongoing controversy over the role of the BBB in uptake of targeted therapeutic agents.⁵ As expected, we were able to show a decrease in 2-HG concentrations from baseline to follow-up scans in the LNT-229 IDH1R132H tumor model during therapy.⁴ Improved SNR for human studies might be achieved at 7T.

Conclusion

In vivo detection of BAY 1436032 using ¹⁹F MRS in an animal model is technically challenging, but feasible.

Acknowledgements

No acknowledgement found.

References

1. Yan H, Parsons DW, Jin G, et al. IDH1 and IDH2 mutations in gliomas. N Engl J Med. 2009;360(8):765-773. doi:10.1056/NEJMoa0808710.
2. Bleeker FE, Lamba S, Leenstra S, et al. IDH1 mutations at residue p.R132 (IDH1(R132)) occur frequently in high-grade gliomas but not in other solid tumors. Hum Mutat. 2009;30(1):7-11. doi:10.1002/humu.20937.
3. Pusch S, Krausert S, Fischer V, et al. Pan-mutant IDH1 inhibitor BAY 1436032 for effective treatment of IDH1 mutant astrocytoma in vivo. Acta Neuropathol. 2017;133(4):629-644. doi:10.1007/s00401-017-1677-y.
4. Bayer AG. Investigator's Brochure, BAY 1436032, Version 3.0, Date 16.04.2018: Identity of Investigational Product: BAY 1436032 Indication: Cancer - mIDH1-R132X mutant malignancies; 2018.
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