Quantitative measurements of GDF-8 using immunoaffinity LC-MS/MS

Area of Science:

• Biochemistry
• Analytical Chemistry
• Pharmacology

Background:

• Growth and differentiation factor 8 (GDF-8) is a key regulator of skeletal muscle mass.
• Muscle loss is associated with various diseases, making GDF-8 a therapeutic target.
• Accurate measurement of GDF-8 is crucial for clinical and translational studies of GDF-8 inhibitors.

Purpose of the Study:

• To develop and validate a specific and sensitive assay for quantifying GDF-8 in serum.
• To enable investigations into the role of GDF-8 in muscle mass regulation.
• To support the development of GDF-8 inhibitors for treating muscle-wasting conditions.

Main Methods:

• Development of an immunoaffinity liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay.
• Offline enrichment of GDF-8 using an anti-GDF-8 antibody and magnetic bead isolation.
• Quantification using 2D nanoflow LC-MS/MS after trypsin digestion.

Main Results:

• The assay was qualified in human serum with a lower limit of quantification (LOQ) of 1.0 ng/mL for intact GDF-8.
• GDF-8 levels were successfully quantified in serum from humans (juvenile and adult) and preclinical species (mouse, rat, cynomolgus monkey).
• The assay demonstrated increased total GDF-8 in cynomolgus monkey serum after administration of an anti-GDF-8 antibody therapeutic.

Conclusions:

• A specific and sensitive LC-MS/MS method for GDF-8 measurement in serum was successfully developed and validated.
• This assay enables confident quantification of GDF-8 across human and preclinical species.
• The ability to accurately measure GDF-8 levels will enhance understanding of its association with muscle mass changes.