Background:⁶⁸Ga-NOTA-WL12 is a peptide-based positron emission tomography (PET) imaging agent. We conducted a first-in-human study of ⁶⁸Ga-NOTA-WL12 for PET to study the in vivo biodistribution, metabolism, radiation dosimetry, safety, and potential for quantifying programmed death ligand-1 (PD-L1) expression levels in advanced non-small cell lung cancer (NSCLC) patients. Methods: In vitro assessment of the relationship between PD-L1 expression and cellular uptake of ⁶⁸Ga-NOTA-WL12 was performed, followed by in vivo evaluation of ⁶⁸Ga-NOTA-WL12 uptake in animal models. Nine NSCLC patients with positive PD-L1 expression in lesions were enrolled. ⁶⁸Ga-NOTA-WL12 and paired ¹⁸F-FDG PET/CT imaging were performed. The patients were monitored for adverse events. The uptake (SUV/L and kBq/mL) values of tumors and normal organs were obtained. The biodistribution, radiation dosimetry and relationship of tumor uptake and PD-L1 expression were then evaluated. Follow-up ¹⁸F-FDG PET/CT was performed in patients who had undergone treatment with a combination of pembrolizumab with chemotherapy. Results: The cellular and animal experiments demonstrated that PD-L1 expression correlated with the uptake of ⁶⁸Ga-NOTA-WL12, and PD-L1-positive tumors exhibited high uptake of ⁶⁸Ga-NOTA-WL12. Clinical research showed that ⁶⁸Ga-NOTA-WL12 PET imaging is safe with acceptable radiation dosimetry. Physiological tracer uptake was mainly visible in the liver, spleen, small intestine and kidney. High contrast of tumors was observed, particularly in the lungs with a T/lung ratio of 4.45 ±1.89 at 1 h. One hour was a suitable timepoint for image acquisition because no significant differences were noted in the tumor-to-background ratios between 1 and 2 h. Additionally, a strong relationship was found between the tumor uptake (SUVpeak) and PD-L1 immunohistochemistry results (r = 0.9349; P = 0.002). ⁶⁸Ga-NOTA-WL12 uptake before therapy might indicate the therapeutic efficacy of pembrolizumab plus chemotherapy combination treatment. Conclusion: Our first-in-human findings demonstrate the safety and feasibility of ⁶⁸Ga-NOTA-WL12 for noninvasive in vivo detection of the tumor PD-L1 expression levels, indicating potential benefits for clinical PD-L1 therapy.

Purpose: Nectin-4 is an emerging biomarker for cancer diagnosis and therapy. Recently, enfortumab vedotin (EV) was approved by the FDA as the first nectin-4 targeting antibody–drug conjugate for treating advanced urothelial carcinoma (UC). A PET imaging method to noninvasively quantify nectin-4 expression level would potentially help to select patients most likely to respond to EV and predict the response. Experimental Design: In this study, we designed a bicyclic peptide-based nectin-4 targeting radiotracer 68Ga-N188. Initially, we performed preclinical evaluations of 68Ga-N188 in UC cell lines and xenograft mouse models. Next, we performed the translational study in healthy volunteers and a pilot cohort of patients with advanced UC on uEXPLORER total-body PET/CT. Results: In the preclinical study, 68Ga-N188 showed high affinity to nectin-4, specific uptake in a nectin-4(+) xenograft mouse model, and suitable pharmacokinetic and safety profiles. In the translational study, 2 healthy volunteers and 14 patients with advanced UC were enrolled. The pharmacokinetic profile was determined for 68Ga-N188, and the nectin-4 relative expression level in different organs was quantitatively imaged. Conclusions: A clear correlation between PET SUV value and nectin-4 expression was observed, supporting the application of 68Ga-N188 PET as a companion diagnostic tool for optimizing treatments that target nectin-4. See related commentary by Jiang et al., p. 3259

Accurately identifying patients who respond to immunotherapy remains clinically challenging. A noninvasive method that can longitudinally capture information about immune cell function and assist in the early assessment of tumor responses is highly desirable for precision immunotherapy. Here, we show that PET imaging using a granzyme B–targeted radiotracer named ⁶⁸Ga-grazytracer, could noninvasively and effectively predict tumor responses to immune checkpoint inhibitors and adoptive T cell transfer therapy in multiple tumor models. ⁶⁸Ga-grazytracer was designed and selected from several radiotracers based on non-aldehyde peptidomimetics, and exhibited excellent in vivo metabolic stability and favorable targeting efficiency to granzyme B secreted by effector CD8⁺ T cells during immune responses. ⁶⁸Ga-grazytracer permitted more sensitive discrimination of responders and nonresponders than did ¹⁸F-fluorodeoxyglucose, distinguishing between tumor pseudoprogression and true progression upon immune checkpoint blockade therapy in mouse models with varying immunogenicity. In a preliminary clinical trial with 5 patients, no adverse events were observed after ⁶⁸Ga-grazytracer injection, and clinical responses in cancer patients undergoing immunotherapy were favorably correlated with ⁶⁸Ga-grazytracer PET results. These results highlight the potential of ⁶⁸Ga-grazytracer PET to enhance the clinical effectiveness of granzyme B secretion–related immunotherapies by supporting early response assessment and precise patient stratification in a noninvasive and longitudinal manner.

The overexpression of integrin α_vβ₆ in pancreatic cancer makes it a promising target for noninvasive PET imaging. However, currently, most integrin α_vβ₆–targeting radiotracers are based on linear peptides, which are quickly degraded in the serum by proteinases. Herein, we aimed to develop and assess a ⁶⁸Ga-labeled integrin α_vβ₆–targeting cyclic peptide (⁶⁸Ga-cycratide) for PET imaging of pancreatic cancer. Methods:⁶⁸Ga-cycratide was prepared, and its PET imaging profile was compared with that of the linear peptide (⁶⁸Ga-linear-pep) in an integrin α_vβ₆–positive BxPC-3 human pancreatic cancer mouse model. Five healthy volunteers (2 women and 3 men) underwent whole-body PET/CT imaging after injection of ⁶⁸Ga-cycratide, and biodistribution and dosimetry were calculated. PET/CT imaging of 2 patients was performed to investigate the potential role of ⁶⁸Ga-cycratide in pancreatic cancer diagnosis and treatment monitoring. Results:⁶⁸Ga-cycratide exhibited significantly higher tumor uptake than did ⁶⁸Ga-linear-pep in BxPC-3 tumor–bearing mice, owing—at least in part—to markedly improved in vivo stability. ⁶⁸Ga-cycratide could sensitively detect the pancreatic cancer lesions in an orthotopic mouse model and was well tolerated in all healthy volunteers. Preliminary PET/CT imaging in patients with pancreatic cancer demonstrated that ⁶⁸Ga-cycratide was comparable to ¹⁸F-FDG for diagnostic imaging and postsurgery tumor relapse monitoring. Conclusion:⁶⁸Ga-cycratide is an integrin α_vβ₆–specific PET radiotracer with favorable pharmacokinetics and a favorable dosimetry profile. ⁶⁸Ga-cycratide is expected to provide an effective noninvasive PET strategy for pancreatic cancer lesion detection and therapy response monitoring.

To tackle the clinical challenge of noninvasively assessing immunotherapy efficacy in patients, here we used positron emission tomography (PET) with ⁶⁸Ga-grazytracer, which targets granzyme B, a crucial effector molecule secreted by activated CD8⁺ T cells. In this phase 1/2 clinical trial (NCT05000372) involving a diverse cohort of 24 patients with solid tumors and lymphomas who received immunotherapies, including immune checkpoint inhibitors (either alone or with chemotherapies) and chimeric antigen receptor-T cell therapy, we examined the in vivo behaviors of ⁶⁸Ga-grazytracer. Primary endpoints were safety, biodistribution, granzyme B specificity, and the predictive utility of ⁶⁸Ga-grazytracer, while secondary endpoint was the relationship between ⁶⁸Ga-grazytracer uptake and tumor immune phenotype. ⁶⁸Ga-grazytracer exhibited a safe profile and specifically targeted granzyme B in patients. ⁶⁸Ga-grazytracer PET showed superior predictive value for short-term prognosis and progression-free survival than those of conventional assessment criteria, including RECIST 1.1 and PERCIST. Moreover, the uptake of ⁶⁸Ga-grazytracer in tumors was significantly higher in those with a “non-desert” immune phenotype than those with an immune “desert” phenotype, thereby meeting the primary and secondary endpoints of this trial. Collectively, we successfully visualized CD8⁺ T cell effector function in humans using ⁶⁸Ga-grazytracer PET, offering insights for enhancing immunotherapy assessment, patient stratification and treatment planning.