이재승 (서울대학교 의과대학)

조주연 (서울대학교 의과대학)

이재승 (서울대학교 의과대학)

장연서 (서울대학교 의과대학)

조주연 (서울대학교 의과대학)

Sterols are an important class of lipids, and play a variety of physiological and pathological processes. However, comprehensive sterol profiling remains a significant challenge, particularly in the identification of unknown sterols (e.g., isomer, isobar) and their low abundances in biological matrices. Herein, we present a targeted LC-MS/MS strategy for the quantitative analysis of sterols using multi-dimensional characterization and picolinyl derivatization. This strategy involves sterol identification using experimental library, followed by quantification via reversed-phase liquid chromatography coupled with parallel reaction monitoring (PRM) mode. The experimental library is curated incorporating multi-dimensional properties such as accurate m/z, isotope pattern, retention time, and co-eluted peak profiles between MS1 and MS2. The strategy enabled to identify a total of 16 sterols in human plasma (NIST SRM 1950), and 11 sterols in the MDA-MB-231 cell.

The strategy was further applied to radiation-resistant triple negative breast cancer (RTR-TNBC) study. RTR-TNBC showed an increase in lanosterol, a cholesterol synthesis precursor, and an accumulation of 27-hyroxycholesterol (27-OHC), an oxidized sterol, compared to TNBC. 27-OHC is known to promote cancer cell migration, invasion, and metastasis via Cytochrome P450 family 27 subfamily A member 1 (CYP27A1) and estrogen receptor β (ERβ). These findings suggest that alterations in sterol metabolism associated with radiation-induced cancer resistance may serve as a potential therapeutic target.

In summary, we proposed a strategy for the multidimensional characterization of different sterol species using LC-MS-based derivatization. This strategy enables comprehensive sterol profiling and offers molecular insights into many physiological and pathological context in biology.