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Parallel, High-Quality Proteomic and Targeted Metabolomic Quantification Using Laser Capture Microdissected Tissues.

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This study presents a new method for simultaneously measuring proteins and metabolites in laser-capture-microdissection samples from prostate cancer patients, improving disease insights.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Proteomics and Metabolomics

Background:

  • Quantitative proteomics and metabolomics of laser-capture-microdissection (LCM) cell populations offer valuable insights into disease mechanisms.
  • However, achieving high-quality quantification from limited LCM samples remains a significant challenge.

Purpose of the Study:

  • To develop and validate a robust, parallel liquid chromatography-mass spectrometry (LC/MS)-based approach for simultaneous global proteomics and targeted metabolomics quantification from the same LCM samples.
  • To apply this method to prostate cancer (PCa) patient biopsies to investigate cellular and regional differences in androgen distribution and identify dysregulated pathways.

Main Methods:

  • An optimized molecular weight cutoff (MWCO) filter separated protein and small-molecule fractions with high recovery.
  • Microscale derivatization and charge-based enrichment enabled ultrasensitive quantification of key androgens.
  • A detergent-cocktail-based sample preparation and the IonStar pipeline ensured reproducible proteomics quantification with minimal missing data.

Main Results:

  • The developed method achieved high-quality, parallel quantification of the proteome and androgens in LCM samples from PCa patients (N=16).
  • Distinct androgen distribution patterns were observed across cancerous and benign epithelial and stromal cells.
  • Evidence of dysregulated pathways involved in tumor-stroma crosstalk in PCa pathology was identified.

Conclusions:

  • This strategy significantly enhances the scope of quantitative omics investigations using LCM samples, facilitating fit-for-purpose method development.
  • The parallel proteomics/metabolomics approach allows precise corroboration of regulatory processes at both molecular levels, reducing batch effects and improving sample utilization.
  • The method, combined with IonStar, is readily adaptable for larger-cohort clinical analyses, offering deeper insights into disease mechanisms.