Lifitegrast Degradation: Products and Pathways
Abstract
1. Introduction
2. Materials and Methods
2.1. General
2.2. Degradations
2.3. Chromatography and Mass Spectrometry
2.4. Nuclear Magnetic Resonance Spectroscopy
2.5. Molecular Modelling
2.6. In Silico Safety Evaluation
3. Results and Discussion
3.1. Initial and Extended Forced Degradation
3.2. Analyzing Oxidative Forced Degradation Mixture
3.3. Extreme Oxidative Degradation
3.4. Targeted Oxidative Degradation
3.5. Comparison of Isolated DP7 to Purchased Analytical Standard of the Same Structure
3.6. Computational DFT Analysis of Degradation Mechanisms
3.7. Proposed Mechanistic Pathways of Oxidative Degradation
3.8. In Silico Toxicology Evaluation of DP5 and DP7
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
| API | Active pharmaceutical ingredient |
| BDE | Bond dissociation energies |
| CREST | Conformer-rotamer ensemble sampling tool |
| DFT | Density functional theory |
| DP | Degradation product |
| EMA | European Medicines Agency |
| ESI | Electrospray ionization |
| FDA | U.S. Food and Drug Administration |
| HMBC | Heteronuclear multiple bond correlation |
| HRMS | High-resolution mass spectrometry |
| ICAM-1 | Intercellular adhesion molecule |
| ICH | The International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use |
| IR | Infrared spectroscopy |
| IRC | Intrinsic reaction coordinate |
| LC | Liquid chromatography |
| LFA-1 | Lymphocyte function-associated antigen 1 |
| LIF | Lifitegrast |
| LOAEL | Lowest observed adverse effect level |
| NMR | Nuclear magnetic resonance |
| NOAEL | No-observed-adverse-effect level |
| PDE | Permitted daily exposure |
| QSAR | Quantitative structure–activity relationship |
| RH | Relative humidity |
| RT | Room temperature |
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| RT/min | Ion | Obs. m/z | Calc. m/z | Error/ppm | Suggested Structure | |
|---|---|---|---|---|---|---|
| LIF | 15.27 | [M-H]− | 613.0597 | 613.0609 | −1.96 | ![]() |
| [M+H]+ | 615.0785 | 615.0754 | 5.04 | |||
| DP7 | 14.31 | [M-H]− | 629.0589 | 629.0558 | 4.93 | ![]() |
| DP3 | 13.86 | [M-H]− | 617.0581 | 617.0558 | 3.73 | ![]() |
| [M+H]+ | 619.0711 | 619.0703 | 1.29 | |||
| DP4 | 12.06 | [M-H]− | 605.0543 | 605.0558 | −2.48 | ![]() |
| [M+H]+ | 607.0735 | 607.0703 | 5.27 | |||
| DP5 | 11.30 | [M-H]− | 633.0549 | 633.0507 | 6.63 | ![]() |
| [M+H]+ | 635.0682 | 635.0652 | 4.72 | |||
| DP8 | 10.51 | [M-H]− | 483.0208 | 483.0190 | 3.73 | ![]() |
| [M+H]+ | 485.0357 | 485.0335 | 4.54 | |||
| DP1 | 8.66 | [M-H]− | 469.0383 | 469.0397 | −2.98 | ![]() |
| [M+H]+ | 471.0561 | 471.0543 | 3.82 |
| Structure | Bond | BDE | Bond | BDE | Bond | BDE |
|---|---|---|---|---|---|---|
![]() | N1–C40 | 60.1 | N15–C16 | 66.6 | C23–H | 103.1 |
| C2–H | 81.7 | C16–H | 75.6 | C30–H | 95.1 | |
| C3–H | 76.4 | C16–C24 | 48.3 | C32–H | 112.6 | |
| C5–Cl | 77.1 | C16–C17 | 37.4 | C33–H | 112.1 | |
| C6–C12 | 76.8 | C17–H | 77.9 | C35–H | 104.8 | |
| C7–Cl | 78.7 | C17–C18 | 78.2 | C36–H | 103.5 | |
| C8–H | 105.8 | C19–H | 103.7 | C37–C40 | 76.6 | |
| C10–H | 70.4 | C20–S | 57.2 | C38–H | 106.3 | |
| C12–N15 | 73.3 | C21–H | 104.6 | |||
| N15–H | 98.5 | C22–H | 103.6 |
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Štefan, L.; Sušanj, I.; Buljević, J.; Roje, M.; Jurin, M.; Buljan, A.; Rinkovec, T.; Vianello, R.; Pocrnić, M.; Galić, N.; et al. Lifitegrast Degradation: Products and Pathways. Pharmaceutics 2025, 17, 1299. https://doi.org/10.3390/pharmaceutics17101299
Štefan L, Sušanj I, Buljević J, Roje M, Jurin M, Buljan A, Rinkovec T, Vianello R, Pocrnić M, Galić N, et al. Lifitegrast Degradation: Products and Pathways. Pharmaceutics. 2025; 17(10):1299. https://doi.org/10.3390/pharmaceutics17101299
Chicago/Turabian StyleŠtefan, Leo, Ivan Sušanj, Jadranka Buljević, Marin Roje, Mladenka Jurin, Anđela Buljan, Tamara Rinkovec, Robert Vianello, Marijana Pocrnić, Nives Galić, and et al. 2025. "Lifitegrast Degradation: Products and Pathways" Pharmaceutics 17, no. 10: 1299. https://doi.org/10.3390/pharmaceutics17101299
APA StyleŠtefan, L., Sušanj, I., Buljević, J., Roje, M., Jurin, M., Buljan, A., Rinkovec, T., Vianello, R., Pocrnić, M., Galić, N., & Čikoš, A. (2025). Lifitegrast Degradation: Products and Pathways. Pharmaceutics, 17(10), 1299. https://doi.org/10.3390/pharmaceutics17101299









