Quantitative assessment of human serum transferrin receptor in breast cancer patients pre- and post-chemotherapy using peptide immunoaffinity enrichment coupled with targeted proteomics.

sTfR, a soluble form of transferrin receptor in serum, has been suggested as an indicator of bone marrow failure in breast cancer patients receiving chemotherapy. However, intensive chemotherapy could also cause a reduction of sTfR to a level below the LOQ of most assays. An advanced liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based targeted proteomics assay coupled with peptide immunoaffinity enrichment (SISCAPA) was developed and validated for monitoring sTfR. Tryptic peptide 681VEYHFLSPYVSPK693 was selected as a surrogate analyte for quantification. High-abundant proteins were first removed from serum, followed by SISCAPA that was effective in surrogate peptide enrichment and sensitivity enhancement. The resulting LOQ can achieve 100ng/ml (~10-fold increase). Then, sTfR levels in breast cancer patients pre- and post-chemotherapy, and healthy volunteers were accurately quantified as 1.77±0.53μg/ml, 0.98±0.26μg/ml and 1.66±0.50μg/ml, respectively, using a standard addition method. While there was no evidence for a difference between patients and healthy volunteers, differential levels of sTfR pre- and post-chemotherapy were obtained. Comparison between SISCAPA-targeted proteomics and ELISA indicated that the former approach provided a lower value of sTfR. SISCAPA-targeted proteomics may allow the quantification of low-abundant proteins in a more accurate manner.