LC-MS/MS method for quantifying the HIV-1 broadly neutralizing antibody PGT 121.414.LS in human serum

Area of Science:

• Immunology
• Analytical Chemistry
• Pharmacology

Background:

• Accurate quantitation of human immunodeficiency virus-1 (HIV-1) broadly neutralizing antibodies (bNAbs) in serum is essential for clinical trials.
• Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is emerging as a viable alternative to ligand-binding assays for therapeutic antibody measurement.

Purpose of the Study:

• To validate a targeted LC-MS/MS method for quantifying the bNAb PGT 121.414.LS in human serum.
• To establish a reliable assay for pharmacokinetic and pharmacodynamic studies of PGT 121.414.LS in HIV-1 treatment and prevention.

Main Methods:

• Nonspecific purification followed by targeted LC-MS/MS analysis using Orbitrap and triple quadrupole mass spectrometry.
• Validation included linearity, reproducibility, accuracy, matrix effects, dilutional integrity, and stability assessments.
• Ammonium sulfate precipitation was used for purification, with stable isotope-labeled peptides as internal standards.

Main Results:

• The validated method demonstrated linear calibration curves from 10-500 μg/mL with a lower limit of quantitation (LLOQ) of 10 μg/mL.
• Quality control samples met ICH M10 acceptance criteria, indicating high accuracy and reproducibility.
• The assay showed minimal carryover, negligible crosstalk, and was assessed for matrix effects, stability, and dilutional integrity.

Conclusions:

• The developed LC-MS/MS assay accurately quantifies PGT 121.414.LS in human serum within the expected therapeutic concentration range.
• This method provides a robust tool for monitoring PGT 121.414.LS levels in clinical studies for HIV-1 prevention and treatment.
• The validated assay supports the clinical development of PGT 121.414.LS by enabling precise pharmacokinetic and pharmacodynamic assessments.