Increased sensitivity of detection of RASSF1A and GSTP1 DNA fragments in serum of prostate cancer patients: Optimisation of diagnostics using OBBPA-ddPCR

Markus Friedemann, Friederike Horn, Katharina Gutewort, Lars Tautz, Carsten Jandeck, Nicole Bechmann, Olga Sukocheva, Manfred P. Wirth, Susanne Fuessel, Mario Menschikowski

Research output: Contribution to journalArticlepeer-review

Abstract

Identification of aberrant DNA methylation is a promising tool in prostate cancer (PCa) diagnosis and treatment. In this study, we evaluated a two-step method named optimised bias-based preamplification followed by digital PCR (OBBPA-dPCR). The method was used to identify promoter hypermethylation of 2 tumour suppressor genes RASSF1A and GSTP1 in the circulating cell-free DNA (cfDNA) from serum samples of PCa patients (n = 75), benign prostatic hyperplasia (BPH, n = 58), and healthy individuals (controls, n = 155). The PCa cohort was further subdivided into subgroups comprising (I) patients with Gleason Scores (GS) ≤ 7 (n = 55), (II) GS ≥ 8 (n = 10), and (III) patients with metastatic PCa diagnosis (n = 10). We found that RASSF1A methylation levels were significantly increased in all 3 PCa subgroups compared to the controls and BPH cohorts (p <0.01 for all comparisons). Fractional abundances of methylated GSTP1 DNA fragments were significantly increased in subgroup III of metastatic PCa patients (p <0.001). RASSF1A methylation analysis was found to be beneficial as a complementary biomarker where further diagnostic validation is most crucial. In combination with free PSA, RASSF1A methylation status helps to identify PCa patients with GS ≥ 8 and grey-zone total PSA values between 2–10 ng/mL. In our study, PCR biases between 80–90% were sufficient to detect minute amounts of tumour DNA with high signal-to-noise ratios as well as high analytical sensitivity and specificity. Both RASSF1A and GSTP1 exhibited strongly increased DNA methylation levels in all metastatic PCa patients. Our data indicates a superior sensitivity of epigenetic biomarker analyses in early detection of PCa metastases that should also help to improve PCa therapy.

Original languageEnglish
Article number4459
Number of pages16
JournalCancers
Volume13
Issue number17
DOIs
Publication statusPublished - 1 Sep 2021

Keywords

  • Cancer diagnostics
  • Cell-free tumour DNA
  • Circulating cell-free DNA (cfDNA)
  • Digital PCR
  • DNA methylation
  • Liquid biopsy
  • Prostate cancer

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