Preclinical evaluation of PSMA expression in response to androgen receptor blockade for theranostics in prostate cancer

Abstract

Background: Prostate-specific membrane antigen (PSMA)-directed radioligand therapy (RLT) is a promising yet not curative approach in castration-resistant (CR) prostate cancer (PC). Rational combination therapies may improve treatment efficacy. Here, we explored the effect of androgen receptor blockade (ARB) on PSMA expression visualized by PET and its potential additive effect when combined with 177Lu-PSMA RLT in a mouse model of prostate cancer.

Methods: Mice bearing human CRPC (C4-2 cells) xenografts were treated with 10 mg/kg enzalutamide (ENZ), with 50 mg/kg bicalutamide (BIC), or vehicle (control) for 21 days. PSMA expression was evaluated by 68Ga-PSMA11 PET/CT and quantified by flow cytometry of tumor fine needle aspirations before treatment and on days 23, 29, 34, and 39 post-therapy induction. For the RLT combination approach, mice bearing C4-2 tumors were treated with 10 mg/kg ENZ or vehicle for 21 days before receiving either 15 MBq (84 GBq/μmol) 177Lu-PSMA617 or vehicle. DNA damage was assessed as phospho-γH2A.X foci in tumor biopsies. Reduction of tumor volume on CT and survival were used as study endpoints.

Results: Tumor growth was delayed by ARB while 68Ga-PSMA11 uptake increased up to 2.3-fold over time when compared to controls. ABR-induced upregulation of PSMA expression was confirmed by flow cytometry. Phospho-γH2A.X levels increased 1.8- and 3.4-fold at 48 h in response to single treatment ENZ or RLT and ENZ+RLT, respectively. Despite significantly greater DNA damage and persistent increase of PSMA expression at the time of RLT, no additional tumor growth retardation was observed in the ENZ+RLT group (vs. RLT only, p = 0.372 at day 81). Median survival did not improve significantly when ENZ was combined with RLT.

Conclusion: ARB-mediated increases in PSMA expression in PC xenografts were evident by 68Ga-PSMA11 PET imaging and flow cytometry. 177Lu-PSMA617 effectively decreased C4-2 tumor size. However, while pre-treatment with ARB increased DNA damage significantly, it did not result in synergistic effects when combined with RLT.

Keywords: 68Ga-PSMA PET/CT; Androgen receptor blockade; PSMA; Prostate cancer; Radioligand therapy.

Conflict of interest statement

Ethics approval

All animal studies were approved by the UCLA Animal Research Committee (ARC; # 2005-090).

Consent for publication

Not applicable.

Competing interests

JCz and CR are co-founders and hold equity in both Sofie Biosciences and Trethera Therapeutics. Intellectual property has been patented by the University of California and has been licensed to Sofie Biosciences and Trethera Therapeutics. No other potential conflict of interest relevant to this article was reported.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1

Experimental design. a Investigation of ARB (ENZ vs. bicalutamide (BIC) vs. vehicle)-mediated PSMA upregulation in C4-2 xenografts. b Testing efficacy of pre-treatment with ENZ before PSMA-RLT in C4-2 tumors. DNA damage was assessed 4 h, 2 and 4 days post RLT by flow cytometric quantification of phospho-γH2A.X in FNA tumor biopsies

Fig. 2

68Ga-PSMA11 PET/CT. C4-2 tumor-bearing mice were imaged with 68Ga-PSMA11 PET/CT on days 0, 23, 29, 34, and 39 post start of therapy with vehicle (top row), BIC (middle), or ENZ (bottom row). Representative PET/CT images of 1 mouse per treatment group are shown

Fig. 3

ARB increases PSMA at the tumor cell surface. Tumor volume of C4-2 (a) tumors was determined by CT (ENZ vs. vehicle, p = 0.05; BIC vs. vehicle, p = 0.02). 68Ga-PSMA11 PET signal (%IAmean/g) in C4-2 (b) tumors (ENZ vs. vehicle, p = 0.02; BIC vs. vehicle, p = n.s.). PSMA mean fluorescent intensity (MFI) is shown for each group (ENZ vs. vehicle, p = 0.03; BIC vs. vehicle, p = n.s.) (c). The fold-change during ARB treatment, related to the pre-treatment volume is shown. Data are shown as mean ± SD. Asterisks indicate significance

Fig. 4

Pre-treatment with ENZ increases 177Lu-PSMA617-induced DNA damage. Phospho-γH2A.X levels were flow cytometrically quantified in FNA tumor biopsies 4 h, 48 h, and 96 h following administration of 177Lu-PSMA617 or vehicle, respectively. The fold-change in phospho-γH2A.X compared to baseline (day before RLT) is shown as mean ± SD. Asterisks indicate significance in ENZ+RLT-treated tumors compared to baseline (ENZ+RLT: 0 vs. 4 h, p = 0.003; 0 vs. 48 h, p < 0.0001)

Fig. 5

Efficacy of combined ARB and 177Lu-PSMA617 radioligand therapy. Tumor volume was determined by CT. The fold-change in tumor volume after RLT (day 1) administration related to the tumor volume immediately prior to RLT (day 0) is shown. The last dose of ENZ was given at day 0. Data are represented as mean ± SD. a Tumor volume change in all groups. Asterisks indicate significance in vehicle and ENZ groups vs. RLT and ENZ+RLT groups, respectively (****p < 0.0001; **p < 0.01). b Tumor volume change in the RLT and ENZ+RLT groups (p = not significant). c Kaplan-Meyer plot for survival