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. 2025 Mar 17;26(6):2686.
doi: 10.3390/ijms26062686.

Tackling Prostate Cancer with Theranostic E5B9-Bombesin Target Modules (TMs): From Imaging to Treatment with UniCAR T-Cells

Liliana R Loureiro  1 , Susan Pike  2   3 , Melinda Wuest  2   3 , Cody N Bergman  2 , Kira R JØrgensen  2 , Ralf Bergmann  4 , Anja Feldmann  1   5   6   7 , Frank Wuest  2   3 , Michael Bachmann  1   5   6   7
Affiliations

Tackling Prostate Cancer with Theranostic E5B9-Bombesin Target Modules (TMs): From Imaging to Treatment with UniCAR T-Cells

Liliana R Loureiro et al. Int J Mol Sci. .

Abstract

Target modules (TMs), intermediate molecules required for UniCAR T-cell therapy, are promising molecules for immunotheranostic approaches. In the current work, we developed TMs containing a monomeric or dimeric form of the antagonist bombesin peptide (BBN2) and assessed their potential for diagnostic imaging using positron emission tomography (PET) as well as immunotherapy in combination with UniCAR T-cells to target and image GRPR expression in prostate cancer. Synthesized monomeric and dimeric BBN2 TMs retained binding to GRPR in vitro. Both BBN2 TMs specifically activated and redirected UniCAR T-cells to eradicate PC3 and LNCaP cancer cells with high efficiency and in a comparable manner. UniCAR T-cells retained a non-exhausted memory phenotype favorable to their persistence and fitness. The 68Ga-labeled BBN2 TMs showed proof-of-target towards GRPR in PC3 and LNCaP xenografts with similar uptake profiles for both BBN2 TMs in dynamic PET experiments. Clearance occurred exclusively through renal elimination. A tremendously increased in vivo metabolic stability of the BBN2 TMs was observed compared to their counterparts without E5B9. Both monomeric and dimeric BBN2 TMs represent novel and promising immunotheranostic tools for application in prostate cancer with exceptionally high in vivo metabolic stability.

Keywords: PET; UniCAR T-cell therapy; bombesin; prostate cancer; theranostics.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Schematic representation of the UniCAR system targeting the GRP receptor using the monomeric and dimeric BBN2 TMs (A); peptide structures of the synthesized E5B9-NOTA-BBN2 monomeric and dimeric TMs (B).
Figure 2
Figure 2
Cytotoxicity assessment (A), EC50 determination (B), and pro-inflammatory cytokine profile (C) of UniCAR T-cells redirected by monomeric and dimeric BBN2 TMs against PC3 and LNCaP cancer cells. Data are shown as mean ± SEM. EC50: half-maximum effective concentration. p < 0.01 (**), p < 0.001 (***), and p < 0.0001 (****).
Figure 3
Figure 3
Activation (A), exhaustion (B), and memory (C) phenotyping of UniCAR T-cells redirected by BBN2 TMs to target PC3 or LNCaP cancer cells. Data are shown as mean ± SEM. p < 0.05 (*), p < 0.0001 (****).
Figure 4
Figure 4
In vitro competitive cell binding of E5B9-NOTA-BBN2 monomeric and dimeric TMs in PC3 cells in comparison to reference gastrin-releasing peptide (GRP). Data are shown as mean ± SEM. IC50: half-maximum inhibition concentration.
Figure 5
Figure 5
Representative PET images after injection of 68Ga-labeled BBN2 TMs and 68Ga-NOTA-BBN2 into PC3 tumor-bearing mice at 60 min p.i. (A). Time–activity curves (TACs) for tumor uptake profiles of 68Ga-labeled monomeric and dimeric TMs versus 68Ga-NOTA-BBN2 alone (B) in PC3 tumors and LNCaP tumors. Data are shown as SUVmean and mean ± SEM.
Figure 6
Figure 6
Representative PET images of in vivo blocking of 68Ga-NOTA-E5B9-BBN2 TM in a PC3 tumor-bearing mouse and 68Ga-labeled E5B9-(BBN2)2 TM in a LNCaP tumor-bearing mouse in the absence and presence of 300 µg of Ava-BBN2 (A) and their respective TACs for tumor uptake profiles (B). Data are shown as SUVmean and mean ± SEM.
Figure 7
Figure 7
In vivo metabolic stability of 68Ga-labeled peptides E5B9-NOTA-BBN2 TM, E5B9-NOTA-(BBN2)2 TM, NOTA-(BBN2)2, and NOTA-BBN2 in venous blood plasma. Data are shown as % of the intact peptide after 5 to 60 min p.i. and as mean ± SEM (n = 3). * Data from Richter et al. [30].
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