Blood-derived biomarkers in prostate cancer

Prostate cancer (PCa) is the second most commonly occurring cancer in men, accounting for 14.1% of all male cases, and the fourth most common cancer overall (Sung et al., 2021). There were more than 1.4 million new cases of prostate cancer worldwide in 2020, based on the World Cancer Research Fund International statistics. The five-year relative survival rate is 98%, (Siegel et al., 2020), however, in the case of metastatic PCa, it is only 30% (Ng et al., 2020).

At present, diagnosis of PCa has a complex process and is based on (1) symptoms for example difficulties in urination, loss of bladder control or more frequent urination (2) the presence of blood in the urine or sperm, (3) erectile dysfunction (4) physical examination, including rectal digital examination, ultrasound examination, (5) blood test to measure soluble prostate-specific antigen (PSA), and (6) imaging-guided tissue sample testing, also called needle biopsy. The only frequently used plasma-derived biomarker, PSA has low selectivity to detect PCa and monitor the cancer progression itself, furthermore, PSA testing may cause overdiagnosis and overtreatment (Wang et al., 2020; Koo et al., 2019). Liquid biopsy represents a minimally invasive method to detect and trace cancer. It has emerged as a candidate to substitute an invasive tissue biopsy for a repeatable and accurate sampling of cancer to enable personalized precision medicine (Ignatiadis et al., 2021; Corcoran, 2020). Many studies have demonstrated that liquid biopsy can help in cancer diagnosis and prognosis as well as in monitoring cancer treatment (Crowley et al., 2013; De Rubis et al., 2019; Kilgour et al., 2020). In the context of PCa, next-generation liquid biopsy applications utilizing various kinds of circulating biomarkers could provide additional and complementary information to the PSA test, thereby facilitating early cancer diagnosis, monitoring of disease status, and personalized treatment options. Here we provide an insight into the circulating tumor cells (CTCs), circulating cell-free nucleic acids (cfDNA or cfRNA), and extracellular vesicles (EVs) that have been already identified as potential biomarkers in PCa patients’ peripheral blood.

Diagnosis of prostate cancer using new-generation blood-derived biomarkers

Localized or early-stage prostate cancer presents a very low number of CTCs, nevertheless, there are some published results on the ability of CTC applications in the diagnosis of PCa. CTCs were detected and counted in patients’ whole blood by immunofluorescent imaging and compared to healthy individuals which were found to be CTC-negative. These studies applied 0.9-5.1 mL or 6-12 mL of peripheral blood and it was reported using both label-dependent and label-free CTC enrichment methods. In the case of PCa patients, CTC detection and identification can be highly improved by staining the PSA marker (Nagrath et al., 2007; Ozkumur et al., 2013; Ried et al., 2020).

Cell-free nucleic acids are another interesting part of the blood in the field of biomarker research. The increased methylated level of FOXA1, GSTP1, HOXD3, RARβ2, RASSF1A, SEPT9, SOX1, and higher levels of promoter DNA methylations of MCAM, ERα, and ERβ have been proved from plasma and serum samples (Constancio et al., 2019; Brait et al., 2017). Furthermore, serum circulating miRNAs, including miR-17-3p and miR-1185-2-3p were found to be a potential tool for distinguishing control and PCa cohorts(Urabe et al., 2019).

EVs have a well-preserved intravesicular content protected by a lipid bilayer, therefore exosomes and microvesicles can be the origins of intact cell-free nucleic acids and proteins from cancer cells. The elevated levels of EV-related miR-1246 from serum and miR-10a-5p, miR-29b-3p in plasma-derived EVs were detected by Nanostring and real-time PCR analysis in PCa patients (Bagirath et al., 2018; Worst et al., 2019). Moreover, useful EV-related PSA, PSMA, HSP90, EpCAM, and EGFR1 protein biomarkers were found in PCa patients’ blood using a lab-on-a-disc microfluidic system for EV isolation and ELISA for protein detection (Sunkara et al., 2019).

Prognosis of prostate cancer using next-generation blood-derived biomarkers

A large amount of the studies of circulating prostate cancer cells have focused on prognostic and/or therapeutic applications in advanced stages of the disease. CTC numbers and features from metastatic PCa patients can inform the Oncoteam of the chance of progression-free survival (PFS) and overall survival (OS). In most cases, baseline CTC enumeration is performed by EpCAM-positive enrichment from 7.5 mL whole blood, followed by CTC-specific immunostaining, and microscopic analysis to predict the PFS and/or OS (Kruijff et al., 2019; Heller et al., 2017; Okegawa et al., 2016). Some other research groups identified the prognostic value of WNT5a, AURKA, BMP7, and PSMA expression on CTCs by real-time PCR analysis (Singhal et al., 2018; Nagaya et al., 2020).

Metastatic PCa patient’s circulating cfDNA mutations (TP53, ATM, BRCA1, BRCA2, CHEK2, AR, MYC, BRAF, RB1) have prognostic value to the OS, treatment failure-free survival, and time to androgen deprivation therapy failure. These genetic alterations were detected by New-Generation Sequencing (NGS) method (Sonpavde et al., 2019; Kohli et al., 2020).

A detectable level of androgen receptor splice variant 7 (AR-V7) plasma-derived exosomal mRNA was associated with a shorter time to progression in castration-resistant prostate cancer patients (Joncas et al., 2019).

Precision medicine and treatment response monitoring using next-generation blood biomarkers

In PCa patients the dynamic changes of CTC count and characteristics have been widely adopted as a way to evaluate the treatment response or disease progression. It was demonstrated in metastatic castration-resistant prostate cancer patient cohort (mCRPC), that response to abiraterone acetate (AA) or enzalutamide (ENZ) can be tracked by CTC enumeration during the first 12-13 weeks of treatment (Lorente et al., 2018; Heller et al., 2018) and by AR-V7 detection in CTCs (Armstrong et al., 2019).

In patients with mCRPC plasma-derived EVs AR-V7 level measurement by digital droplet PCR is a useful method for resistance prediction to hormonal therapy, and response monitoring to AA or ENZ (Del Re et al., 2017).

The cfDNA concentration and/or some cfDNA alterations (TP53, PI3K, BRCA2 and ATM defects, etc.) can be a predictive marker of response to AA and or ENZ therapy (Annala et al., 2018; Torquato et al., 2019). Moreover, cfDNA concentration and DNA repair mutations in BRCA2, and PALB2 genes predict the response to Poly (adenosine diphosphate ribose) polymerase (PARP) inhibitor therapy (Goodall et al., 2017). Interestingly, therapy response prediction to pembrolizumab was also investigated by NGS technology using cfDNA microsatellite instability level quantification in the mCRPC patient cohort (Barata et al., 2020).

Apart from blood biomarkers, a large panel of urine and tissue-based potential markers are identified with diagnostic, prognostic and/or predictive values. Some of these have been already approved for clinical use, and some are still in the experimental stage. Detailed reviews are available on these topics: Wu et al., 2017; Fujita and Nonomura, 2018; Zhao et al., 2014; Eggener et al., 2020. We advise consulting these publications for interested readers.