Ultrafast and Predictive Mass Spectrometry-Based Autotaxin Assays for Label-Free Potency Screening
Tom Bretschneider1, Andreas Harald Luippold1, Helmut Romig1
11 Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
Abstract:
Autotaxin (ATX) is a promising drug target for the treatment of several diseases, such as cancer and fibrosis. ATX hydrolyzes lysophosphatidyl choline (LPC) into bioactive lysophosphatidic acid (LPA). The potency of ATX inhibitors can be readily determined by using fluorescence-based LPC derivatives. While such assays are ultra-high throughput, they are prone to false positives compared to assays based on natural LPC. Here we report the development of ultrafast mass spectrometry-based ATX assays enabling the measurement of data points within 13 s, which is 10 times faster than classic liquid chromatography-mass spectrometry. To this end, we set up a novel in vitro and whole-blood assay. We demonstrate that the potencies determined with these assays are in good agreement with the in vivo efficacy and that the whole-blood assay has the best predictive power. This high-throughput label-free approach paired with the translatable data quality is highly attractive for appropriate guidance of medicinal chemists for constructing strong structure-activity relationships.
Insights
We developed ultrafast mass spectrometry assays for autotaxin (ATX) inhibitors, offering a faster and more reliable method than fluorescence assays. These assays accurately predict in vivo efficacy, aiding drug discovery for cancer and fibrosis.
Area of Science:
- Biochemistry
- Pharmacology
- Analytical Chemistry
Background:
- Autotaxin (ATX) is a key enzyme in lysophosphatidic acid (LPA) production, implicated in diseases like cancer and fibrosis.
- Current fluorescence-based assays for ATX inhibitors are high-throughput but susceptible to false positives.
- A need exists for more accurate and efficient methods to assess ATX inhibitor potency.
Purpose of the Study:
- To develop novel, ultrafast mass spectrometry-based assays for autotaxin (ATX).
- To establish reliable in vitro and whole-blood assays for measuring ATX inhibitor potency.
- To validate the predictive power of these assays against in vivo efficacy.
Main Methods:
- Development of ultrafast mass spectrometry assays capable of 13-second measurements.
- Implementation of novel in vitro and whole-blood assay formats.
- Comparison of assay results with established methods and in vivo efficacy data.
Main Results:
- Ultrafast mass spectrometry assays achieved measurements 10 times faster than conventional LC-MS.
- Assay-determined potencies correlated well with in vivo efficacy.
- The whole-blood assay demonstrated the highest predictive power for drug efficacy.
Conclusions:
- Ultrafast, label-free mass spectrometry assays provide a highly efficient and accurate method for ATX inhibitor evaluation.
- These assays offer superior data quality and predictive power compared to fluorescence-based methods.
- The developed assays are valuable tools for guiding medicinal chemistry efforts in drug discovery for ATX-targeted diseases.
