Radionuclide therapy targeting prostate-specific membrane antigen (PSMA) is a promising option for metastatic castration-resistant prostate cancer. Clinical experience using 177Lu or 225Ac has demonstrated encouraging treatment responses; however, responses are not durable. Dual-isotope combinations, or tandem approaches, may improve tolerability while retaining a high tumor dose. In this study, we directly compared α- versus β-particle treatment, as well as a combination thereof, at different stages of disease in a murine model of disseminated prostate cancer. Methods: First, to determine comparable injected activities from 177Lu- and 225Ac-PSMA-617, ex vivo biodistribution studies were performed at 5 time points after treatment of C4-2 subcutaneous tumor-bearing NSG mice. To establish a more representative model of metastatic prostate cancer, NSG mice were inoculated with luciferase-expressing C4-2 cells in the left ventricle, leading to disseminated visceral and bone lesions. At either 3 or 5 wk after inoculation, the mice were treated with equivalent tumor dose-depositing activities of 177Lu- or 225Ac-PSMA-617 alone or in combination (35 MBq of 177Lu, 40 kBq of 225Ac, or 17 MBq of 177Lu + 20 kBq 225Ac; 10/group). Disease burden was assessed by weekly bioluminescence imaging. Treatment efficacy was evaluated using whole-body tumor burden and overall survival. Results: The ex vivo biodistribution studies revealed that 35 MBq of 177Lu and 40 kBq of 225Ac yield equivalent absorbed tumor doses in a subcutaneous C4-2 model. The disease burden of mice treated at 3 wk after inoculation (microscopic disease) with 177Lu was not significantly different from that of untreated mice. However, 225Ac-PSMA-617 both as a single agent and in combination with 177Lu (17 MBq of 177Lu + 20 kBq of 225Ac) were associated with significant whole-body tumor growth retardation and survival benefit (overall survival, 8.3 wk for nontreatment, 9.4 wk for 177Lu, 15.3 wk for 225Ac alone, and 14.1 wk for tandem therapy). When treated at 5 wk after inoculation (macroscopic disease), all treatment groups showed retarded tumor growth and improved survival, with no significant differences between 225Ac alone and administration of half the 225Ac activity in tandem with 177Lu (overall survival, 7.9 wk for nontreatment, 10.3 wk for 177Lu, 14.6 wk for 225Ac alone, and 13.2 wk for tandem therapy). Conclusion: Treatment of a disseminated model of prostate cancer with simultaneous 225Ac- and 177Lu-PSMA-617 results in significantly decreased tumor growth compared with 177Lu, which was ineffective as a single agent against microscopic lesions. Mice treated later in the disease progression and bearing macroscopic, millimeter-sized lesions experienced significant tumor growth retardation and survival benefit in both monoisotopic and tandem regimens of 177Lu and 225Ac. Although the greatest benefits were observed with the single agent 225Ac, the tandem arm experienced no significant difference in disease burden or survival benefit, suggesting that the reduced activity of 225Ac was adequately compensated in the tandem arm. The superior therapeutic efficacy of 225Ac in this model suggests a preference for α-emitters alone, or possibly in combination, in the microscopic disease setting.

Background

PSMA-TO-1 ("Tumor-Optimized-1") is a novel PSMA ligand with longer circulation time than PSMA-617. We compared the biodistribution in subcutaneous tumor-bearing mice of PSMA-TO-1, PSMA-617 and PSMA-11 when labeled with ⁶⁸Ga and ¹⁷⁷Lu, and the survival after treatment with ²²⁵Ac-PSMA-TO-1/-617 in a murine model of disseminated prostate cancer. We also report dosimetry data of ¹⁷⁷Lu-PSMA-TO1/-617 in prostate cancer patients.

Methods

First, PET images of ⁶⁸Ga-PSMA-TO-1/-617/-11 were acquired on consecutive days in three mice bearing subcutaneous C4-2 xenografts. Second, 50 subcutaneous tumor-bearing mice received either 30 MBq of ¹⁷⁷Lu-PSMA-617 or ¹⁷⁷Lu-PSMA-TO-1 and were sacrificed at 1, 4, 24, 48 and 168 h for ex vivo gamma counting and biodistribution. Third, mice bearing disseminated lesions via intracardiac inoculation were treated with either 40 kBq of ²²⁵Ac-PSMA-617, ²²⁵Ac-PSMA-TO-1, or remained untreated and followed for survival. Additionally, 3 metastatic castration-resistant prostate cancer patients received 500 MBq of ¹⁷⁷Lu-PSMA-TO-1 under compassionate use for dosimetry purposes. Planar images with an additional SPECT/CT acquisition were acquired for dosimetry calculations.

Results

Tumor uptake measured by PET imaging of ⁶⁸Ga-labeled agents in mice was highest using PSMA-617, followed by PSMA-TO-1 and PSMA-11. ¹⁷⁷Lu-PSMA tumor uptake measured by ex vivo gamma counting at subsequent time points tended to be greater for PSMA-TO-1 up to 1 week following treatment (p > 0.13 at all time points). This was, however, accompanied by increased kidney uptake and a 26-fold higher kidney dose of PSMA-TO-1 compared with PSMA-617 in mice. Mice treated with a single-cycle ²²⁵Ac-PSMA-TO-1 survived longer than those treated with ²²⁵Ac-PSMA-617 and untreated mice, respectively (17.8, 14.5 and 7.7 weeks, respectively; p < 0.0001). Kidney, salivary gland, bone marrow and mean ± SD tumor dose coefficients (Gy/GBq) for ¹⁷⁷Lu-PSMA-TO-1 in patients #01/#02/#03 were 2.5/2.4/3.0, 1.0/2.5/2.3, 0.14/0.11/0.10 and 0.42 ± 0.03/4.45 ± 0.07/1.8 ± 0.57, respectively.

Conclusions

PSMA-TO-1 tumor uptake tended to be greater than that of PSMA-617 in both preclinical and clinical settings. Mice treated with ²²⁵Ac-PSMA-TO-1 conferred a significant survival benefit compared to ²²⁵Ac-PSMA-617 despite the accompanying increased kidney uptake. In humans, PSMA-TO-1 dosimetry estimates suggest increased tumor absorbed doses; however, the kidneys, salivary glands and bone marrow are also exposed to higher radiation doses. Thus, additional preclinical studies are needed before further clinical use.

Fibroblast activation protein (FAP)-expressing cancer-associated fibroblasts confer treatment resistance and promote metastasis and immunosuppression. Because FAP is overexpressed in many cancers, radiolabeled molecules targeting FAP are studied for their use as pancancer theranostic agents. This study aimed to establish the spectrum of FAP expression across various cancers by immunohistochemistry and to explore whether ⁶⁸Ga FAP inhibitor (FAPi)-46 PET biodistribution faithfully reflects FAP expression from resected cancer and non-cancer specimens. Methods: We conducted a FAP expression screening using immunohistochemistry on a pancancer human tissue microarray (141 patients, 14 different types of cancer) and an interim analysis of a prospective exploratory imaging trial in cancer patients. Volunteer patients underwent 1 whole-body ⁶⁸Ga-FAPi-46 PET/CT scan and, subsequently, surgical resection of their primary tumor or metastasis. ⁶⁸Ga-FAPi-46 PET SUV_max and SUV_mean was correlated with FAP immunohistochemistry score in cancer and tumor-adjacent non-cancer tissues for each patient. Results: FAP was expressed across all 14 cancer types on tissue microarray with variable intensity and frequency, ranging from 25% to 100% (mean, 76.6% ± 25.3%). Strong FAP expression was observed in 50%-100% of cancers of the bile duct, bladder, colon, esophagus, stomach, lung, oropharynx, ovary, and pancreas. Fifteen patients with various cancer types (colorectal [n = 4], head and neck [n = 3], pancreas [n = 2], breast [n = 2], stomach [n = 1], esophagus [n = 2], and uterus [n = 1]) underwent surgery after their ⁶⁸Ga-FAPi-46 PET/CT scan within a mean interval of 16.1 ± 14.4 d. ⁶⁸Ga-FAPi-46 SUVs and immunohistochemistry scores were higher in cancer than in tumor-adjacent non-cancer tissue: mean SUV_max 7.7 versus 1.6 (P < 0.001), mean SUV_mean 6.2 versus 1.0 (P < 0.001), and mean FAP immunohistochemistry score 2.8 versus 0.9 (P < 0.001). FAP immunohistochemistry scores strongly correlated with ⁶⁸Ga-FAPi 46 SUV_max and SUV_mean: r = 0.781 (95% CI, 0.376-0.936; P < 0.001) and r = 0.783 (95% CI, 0.379-0.936; P < 0.001), respectively. Conclusion: In this interim analysis of a prospective exploratory imaging trial, ⁶⁸Ga-FAPi-46 PET biodistribution across multiple cancers strongly correlated with FAP tissue expression. These findings support further exploration of FAPi PET as a pancancer imaging biomarker for FAP expression and as a stratification tool for FAP-targeted therapies.

Purpose

Prostate-specific membrane antigen (PSMA) targeting radioligands deliver radiation to PSMA-expressing cells. However, the relationship between PSMA levels and intralesion heterogeneity of PSMA expression, and cytotoxic radiation by radioligand therapy (RLT) is unknown. Here we investigate RLT efficacy as function of PSMA levels/cell, and the fraction of PSMA⁺ cells in a tumor.

Experimental design

RM1 cells expressing different levels of PSMA (PSMA^-, PSMA⁺, PSMA⁺⁺, PSMA⁺⁺⁺; study 1) or a mix of PSMA⁺ and PSMA^- RM1 (study 2, 4) or PC-3/PC-3-PIP (study 3) cells at various ratios were injected into mice. Mice received ¹⁷⁷Lu- (studies 1-3) or ²²⁵Ac- (study 4) PSMA617. Tumor growth was monitored. Two days post-RLT, tumors were resected in a subset of mice. Radioligand uptake and DNA damage were quantified.

Results

¹⁷⁷Lu-PSMA617 efficacy increased with increasing PSMA levels (study 1) and fractions of PSMA positive cells (studies 2, 3) in both, the RM1 and PC-3-PIP models. In tumors resected 2 days post-RLT, PSMA expression correlated with ¹⁷⁷Lu-PSMA617 uptake and the degree of DNA damage. Compared with ¹⁷⁷Lu-PSMA617, ²²⁵Ac-PSMA617 improved overall antitumor effectiveness and tended to enhance the differences in therapeutic efficacy between experimental groups.

Conclusions

In the current models, both the degree of PSMA expression and the fraction of PSMA⁺ cells correlate with ¹⁷⁷Lu-/²²⁵Ac-PSMA617 tumor uptake and DNA damage, and thus, RLT efficacy. Low or heterogeneous PSMA expression represents a resistance mechanism to RLT.See related commentary by Ravi Kumar and Hofman, p. 2774.