Figure 2 | Glucocorticoid receptor overexpression. The expression of GR is upregulated after inhibition of AR. AMFR mediate the loss of 11β-HSD2 to maintain cortisol concentrations, which leads to Enz resistance. The loss of TLE3 is another reason of GR upregulation. In addition to the signal target inhibitors, GR/AR dual antagonists Z19 suppresses the transcriptional activity of both AR and GR. GR, glucocorticoid receptor; GC, glucocorticoid; p23, the small, acidic co-chaperone; FKBP52, a large peptidyl-proyl cis–trans isomerase immunophilin protein; AMFR, ubiquitin E3-ligase autocrine mobility factor receptor; 11β-HSD2, 11β-hydroxysteroid dehydrogenase-2; TLE3, transducin-like enhancer of split 3; PI3K/AKT, phosphatidylinositol 3-kinase/ protein kinase B; KLF9, Kruppel-like factor 9; PSA, prostate specific antigen.

Accumulating evidence suggested that Wnt signalling plays complex roles in CRPC and has therapeutic effects of AR-targeting agents. The Wnt family consists of 19 cysteines and secreted lipoglycoproteins, which regulate stem cell self-renewal and cell proliferation, migration, and differentiation during embryonic and organ development. Wnt proteins bind to multiple transmembrane frizzled (FZD) and diverse coreceptors, such as low-density lipoprotein receptor (LRP)4, LRP5, and LRP6, inactive tyrosine-protein kinase transmembrane receptor (ROR)1 and ROR2, and tyrosine-protein kinase (RYK), to activate canonical (β-catenin-dependent) and noncanonical (β-catenin-independent) signalling pathways (Murillo-Garzon and Kypta, 2017).

The Wnt/β-catenin pathway is the best-studied canonical Wnt signalling pathway. It has been reported to affect cell proliferation and differentiation and the epithelial-to-mesenchymal transition (EMT) transition in prostate cancer. The Wnt ligands form complexes with FZD receptors and coreceptors, including LRP5 or LRP6. The complex is inhibited after receptor activation, leading to a blockade of the phosphorylation of β-catenin by both casein kinase Iα and glycogen synthase kinase-3β (GSK-3β). β-catenin is a multifunctional protein that regulates cell adhesion and gene activation. A characteristic of canonical Wnt signalling is the stabilization and nuclear localization of β-catenin. Genomic and RNA-seq analyses demonstrate that Wnt /β-catenin signalling is activated in Enz-resistant CRPC, partially due to a reduction in β-TrCP-mediated ubiquitination. Moreover, the connection between AR and β-catenin is stronger in CRPC tissues than it is in normal tissues. The upregulation of stem-like genes upon the activation of the Wnt/β-catenin pathway contributes to Enz resistance by reactivating AR signalling (Zhang, Cheng et al., 2018). β-catenin is interacts with AR and is an AR coactivator, as indicated by β-catenin armadillo repeats directly interacting with the LBD of AR. Sex-determining region Y(Sry)-related high-mobility group (HMG) box 9 (SOX9), a member of the SOX family of HMG DNA-binding domain-containing transcription factors, regulated β-catenin/T cell factor activity and may be upregulated downstream of Wnt/β-catenin signalling. It was induced by Wnt signalling and positively regulated multiple genes of Wnt pathways. Moreover, AR was one of the SOX9-regulated proteins in PCa cells. The SOX9-AR-Wnt/β-catenin signalling axis in PCa led to the synergistic aberrant expression of the target genes involved in cell viability, multiplication, and differentiation, resulting in the early emergence of invasive and aggressive CRPC. Therefore, targeting the SOX9-AR-Wnt/β-catenin signalling axis may provide a potential strategy for Enz-resistant CRPC treatment (Khurana and Sikka, 2019). In addition, PRKAR2B is also a critical oncogenic gene in CRPC, which belongs to the tetrameric enzyme PKA. PRKAR2B is upregulated in CRPC cells, inducing the EMT and activating the Wnt/β-catenin pathway. Further research indicated that miR-200b-3p, miR-200c-3p, and X-box-binding protein 1 were upstream regulators of PRKAR2B in prostate cancer. Notably, miR-200b-3p and miR-200c-3p were strongly downregulated in CRPC cells and negatively correlated with the expression of PRKAR2B. Given the multiple key factors regulating PRKAR2B, they are additional candidates for therapeutic targets for treatment (Sha, Han et al., 2018 , Xia, Han et al., 2020). The small-molecule Wnt/β-catenin inhibitor CWP232291 inhibited β-catenin and AR, inducing endoplasmic reticulum stress and upregulating the pro-apoptotic transcription factor CHOP. It may also be a candidate for CRPC treatment when chemotherapy fails (Pak, Park et al., 2019).

Noncanonical Wnt signalling is divided into planar cell polarity (PCP) and Wnt-Ca²⁺ pathways. Upon Wnt binding to FZD receptors and tyrosine-kinase-like coreceptors, noncanonical Wnt signalling is activated, leading to the recruitment and activation of Dishevelled (DVL). The PCP pathway has two parallel branches related to small GTPases, including Rho and Rac. Rho activates Rho-associated kinase (ROCK), and Rac is linked to c-Jun N-terminal kinase (JNK). Cytoskeletal and transcriptional changes associated with these small GTPases regulate cell adhesion and migration. The Wnt-Ca²⁺ pathway stimulates Ca²⁺release from the endoplasmic reticulum after being activated, thereby promoting the activation of G-proteins, protein kinase C (PKC),and Ca²⁺/calmodulin-dependent kinase type II (CaMKII) (Murillo-Garzon and Kypta, 2017). RNA-seq of single prostate circulating tumour cells (CTCs) indicated the activation of the noncanonical Wnt signalling pathway induced by anti-androgens. Notably, the noncanonical Wnt ligand Wnt5A enhanced the survival of AR-positive LNCaP cells in the presence of Enz (Miyamoto, Zheng et al., 2015). Subsequent research validated that the finding that the expression of Wnt5A, RhoA, and ROCK was increased in Enz-resistant PCa cells. Moreover, the combination of the ROCK inhibitor Y27632 and Enz synergistically inhibited 22RV1‐derived xenograft tumour growth. Additionally, ROCK deletion combined with Enz inhibited the invasion and migration of Enz-resistant PCa cells by suppressing the EMT (Chen, Liu et al., 2020).

In addition to the activation of Wnt signalling pathways, Wnt-activating mutations may affect Enz therapy in CRPC. Since Wnt-activating mutations are present in 10-20% of advanced prostate cancers, a recent study found that metastatic CRPC (mCRPC) patients with Wnt pathway mutations showed poor therapeutic effects after abiraterone or Enz treatment. Even after adjusting for multivariate factors and considering the alterations in other vital tumour suppressor genes, the negative prognostic impact of Wnt pathway mutations persisted. Targeting Wnt alterations may provide another idea for Enz-resistant treatment (Isaacsson Velho, Fu et al., 2020). Lombard et al. took this novel step and explored the role of Wnt-secretion-mediating proteins (Lombard, Liu et al., 2019). RNA-seq and gene set enrichment analysis were utilized to validate the expression profile changes in Enz-resistant C4-2B-MDVR cells compared to that of the parental C4-2B cells. Wntless (WLS) was needed for the secretion of all Wnts and mediated the transport of Wnts. WLS was found to be overexpressed in the resistant cells, regulating Wnt signalling and cellular viability. Furthermore, WLS regulated AR and AR variant expression and downstream signalling. These findings support the idea that targeting Wnt secretion may be a potential strategy for treating Enz-resistant CRPC