Beta-blockers prolong response to androgen deprivation therapy in prostate cancer through modulation of the neuro-immuno-oncology axis

Abstract

Background: The therapeutic impact of beta-blockers (BB), beta-adrenergic receptor antagonists, on prostate cancer remains controversial. The underlying health conditions of BB users complicate the ability to isolate and evaluate the specific effects of these drugs on the tumour cells. This study investigated whether BBs, by inhibiting sympathetic nerve signalling, extended the duration of androgen deprivation therapy (ADT) effectiveness in patients with de novo metastatic hormone sensitive prostate cancer and in prostate cancer xenograft models, while also uncovering the molecular mechanisms involved.

Methods: An analysis was conducted on prospectively collected data from the Cancer Registry of Norway, Norwegian Prescription Database, and Norwegian Cause of Death Registry focusing on patients with de novo metastatic prostate cancer undergoing ADT using the commencement of second-line treatment as the endpoint. In addition, the causal effect of BB treatment was studied in two different hormone-sensitive prostate cancer xenograft mouse models. Prior to treatment, mice were surgically castrated, to mimic ADT, and tumour progression was tracked by measuring serum PSA levels. RNA sequencing was performed on xenografted orthotopic tumours to investigate the underlying mechanisms, utilizing annotation based on human data and protein levels were validated by the Protein Simple Immunoassay. Immune-related effects were evaluated using immunohistochemistry on tumour tissue and measuring neopterin levels, along with 92 analytes, using the OLINK proximity extension assay on serum samples from xenografted mice and prostate cancer patients, both BB users and non-users.

Results: A competitive risk analysis indicated that BB treatment postponed the initiation of second-line treatment in prostate cancer patients on ADT. Additionally, in both prostate cancer xenograft models, BB treatment reduced tumour burden and delayed progression to castration-resistant prostate cancer. Mechanistically, BB treatment suppressed androgen receptor signalling and induced a metabolic shift by up-regulating oxidative phosphorylation transcripts and down-regulating those involved in fatty acid synthesis and the PI3K/AKT/mTOR pathway. Additionally, BB treatment increased serum pro-inflammatory cytokines, such as the IL23/IL17 axis, in both xenografted mice and in patient samples. Enhanced intra-tumoral CD68+ immune cell infiltration was also observed in the tumours.

Conclusion: The data suggest that BB combined with ADT delay the progression to castration-resistant prostate cancer. This may be achieved by influencing androgen receptor activity, adjusting energy metabolism and fostering a pro-inflammatory antitumoral microenvironment.

Keywords: Androgen receptor; Beta-adrenergic receptor; Castration-resistant prostate cancer; Oxidative phosphorylation; PI3K/AKT/mTOR; Pro-inflammatory; SRC.

Fig.1

Cumulative incidence curves for beta-blocker (BB) users and non-users. A The cumulative incidence of initiating second-line treatment, considering all-cause mortality as a competing event, was assessed among 1198 patients diagnosed with de novo metastatic prostate cancer, including 360 who were BB users. B The cumulative incidence of time to prostate cancer specific mortality with other causes of death as the competing event. Shaded areas represent the 95% CI and the risk tables are shown below each plot

Fig. 2

Beta-blockers delayed onset of castration resistant prostate cancer. A LNCaP cells were injected into the prostate in nude Balb/c mice. The mice were castrated after three weeks and subsequently treated with vehicle (control, n = 7), propranolol (BB1/2, n = 6) or propranolol and SR59230 A (BB1/2/3, n = 7) for five weeks. Serum and tumours were collected. B Box plots showing serum PSA in mice treated with vehicle (control), propranolol (BB1/2) or the combination of propranolol and SR59230 A (BB1/2/3) at different time points. C Box plots depicting tumour weight at end of study for castrated LNCaP xenograft mice treated with vehicle (control), propranolol (BB1/2) or the combination of propranolol and SR59230 A (BB1/2/3). Data were statistically analysed using Kruskal–Wallis test with Dunn’s as post-hoc test

Fig. 3

Androgen receptor signalling related transcripts are downregulated in BB-treated tumours. Bulk RNA sequencing from orthotopic LNCaP tumours treated with vehicle (Control, n = 7), propranolol (BB1/2, n = 6) or propranolol and SR50230 A (BB1/2/3, n = 6) was performed at the end of study.Volcano plots showing RNA transcripts differentially expressed between mice treated with propranolol (BB1/2, n=6) (A) relative to vehicle (Control, n=7); propranolol and SR59230A (BB1/2/3, n=6) (B) relative to Control; BB1/2 relative to BB1/2/3 (C). Box plots showing the expression level of selected genes encoding the androgen receptor (AR) (D), the androgen-regulated transcription factor, homeobox transcription factor NKX3-1 (E) and ETS homologous factor EHF (F), (genes encoding key enzymes in fatty acid synthesis, ATP citrate lyase (ACLY) (G), Acetyl-CoA carboxylase alpha (ACACA) (H) and fatty acid synthetase (FASN) (I), and the transcript encoding the proliferation marker Ki67 (MKI67) (J) and genes related to senescence (Senescence signature) (K). FPKM: Fragments Per Kilobase Million. p-values were determined using Kruskal–Wallis test with Dunn’s as post-hoc test. L Illustration showing the central role of acetyl-CoA both in fatty acid synthesis and regulation of histone acetylation. Box plots showing RNA seq data for the histone acetyl transferase p300 (EP300) (M) at end of study from tumours of castrated LNCaP xenograft mice treated with vehicle (control), propranolol (BB1/2) or the combination of propranolol and SR59230 A (BB1/2/3). Box plots showing RNA seq data for the histone lysine methyltransferase SET7D (N) at end of study from tumours of castrated LNCaP xenograft mice treated with vehicle (control), propranolol (BB1/2) or the combination of propranolol and SR59230 A (BB1/2/3). p-values were determined using Kruskal–Wallis test with Dunn’s as post.doc test

Fig. 4

Beta-blocker treatment upregulated OXPHOS pathway genes and downregulated chromatin modelling encoding genes and “Endocrine resistance” annotated transcripts. A Volcano plot showing “Endocrine resistant” annotated genes significantly correlated with tumour weight. The horizontal dashed line indicates p-value 0.05 and vertical dashed lines indicates rho values 0.4 and − 0.4. B Box plots showing the sum of gene expression (FPKM) of RNAseq data of PI3K/AKT MTOR genes significantly correlated with tumour weight (PI3K/AKT/mTOR pathway) at end of study. C Box plots presenting the protein and phosphorylation levels of AKT/pAKT/mTOR/p-mTOR/RPS6KB1 (S6) and protein level of SRC in the control (n = 5) and BB1/2/3 (n = 5) groups. GAPDH was used for normalization. (Raw data is presented in supplementary Fig S4. D Volcano plot showing OXPHOS gene expression levels correlated with tumour weight in LNCaP xenograft tumours treated with vehicle (control, n = 7), propranolol (BB1/2, n = 6) and propranolol in combination with SR59230 A (BB1/2/3, n = 6). The horizontal dashed line indicates p-value 0.05 and vertical dashed lines indicates rho values 0.4 and − 0.4. E Box plots showing the FPKM sum of OXPHOS genes significantly correlated with tumour weight (OXPHOS signature) from mice treated with vehicle (control), propranolol (BB1/2) or the combination of propranolol and SR59230 A (BB1/2/3). The list of genes included in the OXPHOS signature is presented in Supplementary Table S4. The signature is based on RNAseq data from the nine mRNAs showing the highest correlation with tumour weight. p-values were determined using Kruskal–Wallis test with Dunn’s as post-hoc test

Fig. 5

Beta-blocker treatment induces a proinflammatory phenotype. Boxplot showing neopterin level at 3 weeks (A) and 5 weeks (B) and density of CD68+ cells (C) at end of study. D Hierarchical clustering analyses were performed on 92 analytes (OLINK) analysed in serum from castrated LNCaP xenograft mice treated with vehicle (control, n = 7), propranolol (BB1/2, n = 6) and propranolol in combination with SR59230 A (BB1/2/3, n = 6). Mice that developed castration-resistant prostate cancer (CRPC) before termination are denoted by dark green, while those that did not progress to CRPC are indicated in light green. E Volcano plot showing serum analytes (OLINK) correlated with tumour weight in LNCaP xenograft tumours. The horizontal dashed line indicates p-value 0.05 and vertical dashed lines indicates rho values 0.4 and − 0.4. F Volcano plot showing serum analytes differentially expressed between prostate cancer patients using or not using beta-blockers. The horizontal dashed red line indicates p-value 0.05 and black line is p-value 0.1. p-values were determined using Kruskal–Wallis test with Dunn’s as post.doc test

Fig. 6

An illustration depicting the molecular changes linked to beta-blocker treatment of prostate cancer xenograft mice undergoing androgen deprivation therapy. Androgen deprivation therapy (ADT) is recognized for inducing apoptosis and growth arrest in prostate tumour cells. Over time, however, growth arrested cancer cells can reactivate leading to proliferation of castration-resistant prostate cancer (CRPC) cells. Our study showed that beta-blocker (BB) treatment amplified molecular effects typically regulated by ADT, resulting in delayed progression to castration-resistant prostate cancer (CRPC). Beta-blockers (BBs) inhibit the effects of sympathetic nerve activation by acting as antagonists for beta-2 adrenergic receptors (ADRB2) and beta-3 adrenergic receptors (ADRB3). The underlying mechanism involved the activation of the proinflammatory IL23/IL17 axis and reduced expression of genes associated with signalling pathways known to promote CRPC, such as AR- and PI3K/AKT/mTOR components. Consistent with these findings, we noted an increased expression of genes coding for components of oxidative phosphorylation and a decreased expression of genes involved in fatty acid synthesis. Additionally, elevated serum levels of pro-glucagon observed following beta-blocker treatment. Created in BioRender. Taskén, K. (2025) https://BioRender.com/w15f914