Abstract
BackgroundThe availability of serum collections from the Women's Health Initiative (WHI) conjugated equine estrogens (CEE) randomized controlled trial provides an opportunity to test the potential of in-depth quantitative proteomics to uncover changes in the serum proteome related to CEE and to assess their relevance to trial findings, including elevations in the risk of stroke and venous thromboembolism and a reduction in fractures.MethodsFive independent large scale quantitative proteomics analyses were performed, each comparing a set of pooled serum samples collected from 10 subjects, 1 year following initiation of CEE at 0.625 mg/d, relative to their baseline pool. A subset of proteins that exhibited increased levels with CEE by quantitative proteomics was selected for validation studies.ResultsOf 611 proteins quantified based on differential stable isotope labeling, the levels of 116 (19%) were changed after 1 year of CEE (nominal P < 0.05), while 64 of these had estimated false discovery rates <0.05. Most of the changed proteins were not previously known to be affected by CEE and had relevance to processes that included coagulation, metabolism, osteogenesis, inflammation, and blood pressure maintenance. To validate quantitative proteomic data, 14 proteins were selected for ELISA. Findings for ten - IGF1, IGFBP4, IGFBP1, IGFBP2, F10, AHSG, GC, CP, MMP2, and PROZ - were confirmed in the initial set of 50 subjects and further validated in an independent set of 50 additional subjects who received CEE.ConclusionsCEE affected a substantial fraction of the serum proteome, including proteins with relevance to findings from the WHI CEE trial related to cardiovascular disease and fracture.Clinical Trials RegistrationClinicalTrials.gov identifier: NCT00000611
Highlights
The availability of serum collections from the Women’s Health Initiative (WHI) conjugated equine estrogens (CEE) randomized controlled trial provides an opportunity to test the potential of in-depth quantitative proteomics to uncover changes in the serum proteome related to CEE and to assess their relevance to trial findings, including elevations in the risk of stroke and venous thromboembolism and a reduction in fractures
CEE affected a substantial fraction of the serum proteome, including proteins with relevance to findings from the WHI CEE trial related to cardiovascular disease and fracture
In-depth proteomic mass spectrometry (MS) analysis of plasmas obtained from subjects in the WHI hormone replacement therapy trial uncovered 116 proteins (19%) that exhibited quantitative changes 1 year after CEE
Summary
The availability of serum collections from the Women’s Health Initiative (WHI) conjugated equine estrogens (CEE) randomized controlled trial provides an opportunity to test the potential of in-depth quantitative proteomics to uncover changes in the serum proteome related to CEE and to assess their relevance to trial findings, including elevations in the risk of stroke and venous thromboembolism and a reduction in fractures. The Women’s Health Initiative (WHI) randomized, placebo controlled trial of 0.625 mg/d continuous CEE among 10,739 women who were post-hysterectomy did not provide evidence of benefit for CHD, and health benefits and risks appeared to be approximately balanced [4]. Adverse effects observed in the WHI trial include increased incidence of venous thromboembolism and stroke [4,12,13]
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