Abstract
Novel androgen receptor (AR) signaling inhibitors have improved the treatment of castration-resistant prostate cancer (CRPC). Nonetheless, the effect of these drugs is often time-limited and eventually most patients become resistant due to various AR alterations. Although liquid biopsy approaches are powerful tools for early detection of such therapy resistances, most assays investigate only a single resistance mechanism. In combination with the typically low abundance of circulating biomarkers, liquid biopsy assays are therefore informative only in a subset of patients. In this pilot study, we aimed to increase overall sensitivity for tumor-related information by combining three liquid biopsy approaches into a multi-analyte approach. In a cohort of 19 CRPC patients, we (1) enumerated and characterized circulating tumor cells (CTCs) by mRNA-based in situ padlock probe analysis, (2) used RT-qPCR to detect cancer-associated transcripts (e.g., AR and AR-splice variant 7) in lysed whole blood, and (3) conducted shallow whole-genome plasma sequencing to detect AR amplification. Although 44–53% of patient samples were informative for each assay, a combination of all three approaches led to improved diagnostic sensitivity, providing tumor-related information in 89% of patients. Additionally, distinct resistance mechanisms co-occurred in two patients, further reinforcing the implementation of multi-analyte liquid biopsy approaches.
Highlights
Prostate cancer (PC) is one of the most common malignant cancers in men causing more than350,000 deaths worldwide per year [1]
We evaluated the in situ padlock probe assay in cells of four healthy donors captured on the CellCollector, in peripheral blood mononuclear cells (PBMCs) of two healthy donors enriched by density gradient centrifugation of blood samples, as well as in the PC cell lines PC-3 and VCaP
These results demonstrate the high specificity of in situ padlock probes, as 99.23–99.29% of PBMCs did not show any false-positive signals for AR-V7, and 98.91–99.04% of PBMCs did not show any false-positive signals for AR-FL
Summary
Prostate cancer (PC) is one of the most common malignant cancers in men causing more than350,000 deaths worldwide per year [1]. One ofcommon the mosttreatment common malignant cancers indeprivation men causingtherapy more than (AR) signaling theis most involves androgen (ADT). AR-V7, AR-FL, and KLK3 showed background signals in blood of healthy donors and we defined a cut-off value based on the healthy donors’ signals [16]. This background decreases the applicability of this method, as only one patient was positive for AR-V7 based on our cut-off value. The enrichment-free RT-qPCR approach is challenged by a background of blood cell RNA signals. The major advantage is the inexpensive technical equipment needed so it can be performed in parallel to more elaborate liquid biopsy approaches
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