Medizinische Universität Graz - Research portal
Selected Publication:
Bonstingl, L.
Tracking the resistance: liquid biopsy monitoring of drug resistance in metastatic prostate cancer
PhD-Studium (Doctor of Philosophy); Humanmedizin; [ Dissertation ] Medizinische Universitt Graz; 2023. pp.
- Authors Med Uni Graz:
- Bonstingl Lilli
- Advisor:
- Bauernhofer Thomas
- El-Heliebi Amin
- Heitzer Ellen
- Altmetrics:
- Abstract:
- Treatment options for metastatic prostate cancer (PC) are continuously improving. Treatment decisions, however, remain challenging due to the diversity of resistance mechanisms and a lack of reliable tools for patient stratification and resistance monitoring. In this context, liquid biopsy studies analyzing circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) in blood samples yield promising results. Yet, they are often informative only in a subset of patients or fail to capture the big picture. Comprehensive liquid biopsy assays and longitudinal monitoring are imperative to advance our understanding of dynamic resistance mechanisms and the clinical implications of liquid biopsies for treatment decisions. We hypothesize, that multiple resistance mechanisms can be detected by mRNA-based single-cell analysis of CTCs. This thesis aimed at establishing a comprehensive mRNA-based CTC analysis workflow that is intended to be used for longitudinal monitoring in combination with serial ctDNA profiling. For the new CTC analysis workflow, we developed a novel combinatorial dual color (CoDuCo) in situ padlock probe hybridization assay that visualizes transcripts in eleven pseudocolors, we implemented semiautomated image analysis and machine learning-assisted CTC classification, and created an interactive image gallery for easy revision of large datasets. The CoDuCo in situ assay’s high multiplex capacity enabled us to visualize a panel of hematopoietic, mesenchymal, epithelial, prostate-specific, and neuroendocrine transcripts for CTC identification and characterization. When tested on seven patient samples, we captured single CTCs and CTC clusters with epithelial, mesenchymal, neuroendocrine, and mixed phenotypes. We determined the expression levels of treatment-related transcripts, visualized diverse resistance mechanisms, predictive biomarkers and druggable targets, revealed CTC heterogeneity, and tracked dynamic changes in five longitudinally collected samples from a multidrug-resistant metastatic PC patient. In this thesis, we confirmed the feasibility and high informative value of CoDuCo in situ CTC analysis and pointed out technical limitations to be tackled in the future. The clinical relevance of the CoDuCo in situ assay, in combination with other liquid biopsy assays, will be investigated in an observational longitudinal cohort study with 25 patients and >130 blood samples. Based on the data presented in this thesis, we believe that the combination of high multiplex capacity and microscopy-based single-cell analysis is a unique and powerful feature of the CoDuCo in situ assay and holds promise to bring new insights into the dynamic molecular changes associated with drug response and resistance.