Tumor Uptake Research Articles

Enhancing Anti-PD-1 Immunotherapy by Targeting MDSCs via Hepatic Arterial Infusion in Breast Cancer Liver Metastases.

Surgery, chemotherapy, and radiation often have limited utility for advanced metastatic disease in the liver, and despite its promising activity in select cancers, PD-1 blockade therapy similarly has minimal benefit in this setting. Curaxin, CBL0137, is an experimental anti-cancer drug that disrupts the binding of DNA to histones, destabilizes chromatin, and induces Z-DNA formation which may stimulate anti-tumor immune responses. Murine cell lines of colon (CT26) and breast (4T1) cancer were interrogated for survival and CBL0137-associated DNA changes in vitro. Immunocompetent models of liver metastases followed by CBL0137 hepatic arterial infusion (HAI) were used to examine in vivo tumor cell DNA alterations, treatment responses, and the immune contexture associated with CBL0137, both alone and in combination with anti-PD-1 therapy. CBL0137 induced immediate changes to favor tumor cell death in vitro and in vivo with an efficient tumor uptake via the HAI route. Toxicity to CBL0137 was minimal and anti-tumor treatment effects were more efficient with HAI compared to intravenous delivery. Immune effects were pronounced with CBL0137 HAI with concurrent depletion of a specific population of myeloid-derived suppressor cells and maintenance of effector T cell populations. Combination of CBL0137 HAI with PD-1 blockade improved survival in 4T1 tumors but not in CT26 tumors, and therapeutic efficacy relies on the finding of simultaneous and targeted depletion of myeloid-derived suppressor cells and skewing of T cell populations to produce synergy with PD-1 blockade therapy.

Molecular Imaging of Melanoma VEGF-expressing Tumors through [99mTc]Tc-HYNIC-Fab(Bevacizumab).

Angiogenesis is a process that many tumors depend on for growth, development, and metastasis. Vascular endothelial growth factor (VEGF) is one of the major players in tumor angiogenesis in several tumor types, including melanoma. VEGF inhibition is achieved by bevacizumab, a humanized monoclonal antibody that binds with high affinity to VEGF and prevents its function. In order to successfully enable in vivo VEGF expression imaging in a murine melanoma model, we previously labeled bevacizumab with [99mTc]Tc. We observed that this was feasible, but it had prolonged blood circulation and delayed tumor uptake. The aim of this study was to develop a radiolabeled Fab bevacizumab fragment, [99mTc]Tc-HYNICFab( bevacizumab), for non-invasive in vivo VEGF expression molecular imaging. Flow cytometry was used to examine VEGF presence in the murine melanoma cell line (B16-F10). Bevacizumab was digested with papain for six hours at 37°C to produce Fab(bevacizumab), which was then conjugated to NHS-HYNIC-Tfa for radiolabeling with [99mTc]Tc. Stability and binding affinity assays were also evaluated. Biodistribution and single photon emission computed tomography/computed tomography (SPECT/CT) were performed at 1, 3, and 6 h (n = 4) after injection of [99mTc]Tc-HYNIC-Fab(Bevacizumab) in normal and B16-F10 tumor-bearing C57Bl/6J mice. Using flow cytometry, it was shown that the B16-F10 murine melanoma cell line has intracellular VEGF expression. Papain incubation resulted in the complete digestion of bevacizumab with good purity and homogeneity. The radiolabeling yield of [99mTc]Tc-HYNIC-Fab(bevacizumab) was 85.00 ± 6.06%, with a specific activity of 291.87 ± 18.84 MBq/mg (n=3), showing in vitro stability. Binding assays demonstrated significant intracellular in vitro VEGF expression. Fast blood clearance and high kidney and tumor uptake were observed in biodistribution and SPECT/CT studies. We present the development and evaluation of [99mTc]Tc-HYNIC-Fab(bevacizumab), a novel molecular VEGF expression imaging agent that may be used for precision medicine in melanoma and potentially in other VEGF-expressing tumors.

Preliminary Study of Radionuclide-Labeled MerTK-Targeting PET Imaging Agents for the Diagnosis of Melanoma.

MerTK PET imaging holds potential as a promising approach for assessing tumor aggressiveness and monitoring treatment response. In this study, we synthesized a series of 18F- and 68Ga-labeled tracers derived from MerTK inhibitors for detection of MerTK expression. Among the synthesized agents, the dimeric compounds [68Ga]10 and [68Ga]12 demonstrated good in vivo and in vitro stability, high affinities to the MerTK receptor, and good MerTK-targeting specificity. Notably, [68Ga]10 exhibited a tumor uptake of 2.6 ± 0.2%ID/g at 1 h p. i. in B16F10 tumor-bearing mice, nearly tripling the uptake of its monomeric counterpart [68Ga]3. A similar enhancement was observed with [68Ga]12 compared to its monomeric analog [68Ga]6. Additionally, [18F]14 achieved a tumor uptake of 7.6 ± 0.5%ID/g at 2 h p. i., outperforming the previously reported [18F]15. Biodistribution analysis further validated the results, highlighting their potential for clinical investigation.

Synthesis and preclinical evaluation of [18F]AlF-NODA-MP-C6-CTHRSSVVC as a PET tracer for CD163-positive tumor-infiltrating macrophages

Positron emission tomography (PET) can provide information about tumor-associated macrophage (TAM) infiltration, as long as a suitable tracer is available. This study aimed to evaluate the radiolabeled peptide [18F]AlF-NODA-MP-C6-CTHRSSVVC as a potential PET tracer for imaging of the CD163 receptor, which is expressed on M2-type tumor-associated macrophages. The conjugated peptide NODA-MP-C6-CTHRSSVVC was labeled with aluminum [18F]fluoride. Tracer binding and its biodistribution were evaluated in an in vitro binding assay and in healthy BALB/c mice, respectively. In addition, different treatments with cyclophosphamide in tumor-bearing mice were used to assess whether the tracer could detect differences in CD163 expression caused by differential TAM infiltration. After 7 days of treatment, animals were injected with [18F]AlF-NODA-MP-C6-CTHRSSVVC, and a 60-min dynamic PET scan was performed, followed by an ex vivo biodistribution study. [18F]AlF-NODA-MP-C6-CTHRSSVVC was prepared in 23 ± 6 % radiochemical yield and showed approximately 50 % of specific receptor-mediated binding in an in vitro binding assay on human CD163-expressing tissue homogenates. No CD163-mediated binding of [18F]AlF-NODA-MP-C6-CTHRSSVVC was detected by PET under normal physiological conditions in healthy BALB/c mice. On the other hand, CD163-positive xenograft tumors were clearly visualized with PET and a positive correlation was found between CD163 levels and the [18F]AlF-NODA-MP-C6-CTHRSSVVC tumor-to-muscle ratio (TMR) obtained from the PET images (Pearson r = 0.76, p = 0.002). No significant differences in the CD163 protein level and in the tracer uptake between treatment groups were found in the tumors. Taken together, [18F]AlF-NODA-MP-C6-CTHRSSVVC appears a promising candidate PET tracer for M2-type TAM, as it binds specifically to CD163 in vitro and its tumor uptake correlates well with CD163 expression in vivo.

Synthesis and evaluation of a novel BODIPY fluorescent probe targeting integrin αvβ3 for cancer diagnosis.

A series of integrin αvβ3 targeting BODIPY-RGD conjugate were designed and synthesized. Their in vitro and in vivo fluorescence imaging behaviors were investigated. The small molecule compound was designed as an optical imaging near-infrared fluorescent dye which was combined RGD peptide with the meso-amide BODIPYs using succinic moiety as a spacer. The construction alleviated the traditional BODIPY problems including poor water solubility, aggregate caused quench (ACQ) effect, low biocompatibility, etc. In cellular research, BDP-RGD-2 showed rapid, selective uptake in 3 highly expressing integrin αvβ3 cell lines MDA-MB-231, A549, U87MG at different extent rather than an integrin αvβ3 low level expression cell MCF-7. In animal study, fluorescence imaging of U87MG model targeted by BDP-RGD-2 displayed a highest tumor uptake level and T/N ratio up to 6h after tail-intravenous injection, which demonstrated BDP-RGD-2 was a promising probe for tracing integrin αvβ3 overexpressing tumors.

Kinetic Analysis and Metabolism of Poly(Adenosine Diphosphate-Ribose) Polymerase-1-Targeted 18F-Fluorthanatrace PET in Breast Cancer.

The poly(adenosine diphosphate-ribose) polymerase inhibitors (PARPi) have demonstrated efficacy in ovarian, breast, and prostate cancers, but current biomarkers do not consistently predict clinical benefit. 18F-fluorthanatrace (18F-FTT) is an analog to rucaparib, a clinically approved PARPi, and is a candidate biomarker for PARPi response. This study intends to characterize 18F-FTT pharmacokinetics in breast cancer and optimize image timing for clinical trials. A secondary aim is to determine whether 18F-FTT uptake in breast cancer correlates with matched frozen surgical specimens as a reference standard for PARP-1 protein. Methods: Thirty prospectively enrolled women with a new diagnosis of breast cancer were injected with 18F-FTT and imaged dynamically 0-60 min after injection over the chest, with an optional static scan over multiple bed positions starting around 70 min. Kinetic analysis of lesion uptake was performed using blood-pool activity with population radiometabolite corrections. Normal breast and normal muscle reference tissue models were compared with PARP-1 protein expression in 10 patients with available tissue. Plasma radiometabolite concentrations and uptake in tumor and normal muscle were investigated in mouse xenografts. Results: Pharmacokinetics of 18F-FTT were well fit by Logan plot reference region models of reversible binding. However, fits of 2-tissue compartment models assuming negligible metabolite uptake were unstable. Rapid metabolism of 18F-FTT was demonstrated in mice, and similar uptake of radiometabolites was found in tumor xenografts and normal muscle. Tumor 18F-FTT distribution volume ratios relative to normal muscle reference tissue correlated with tissue PARP-1 expression (P < 0.02, n = 10). The tumor-to-normal muscle ratio from a 5-min frame between 50 and 60 min after injection, a potential static scan protocol, closely corresponded to the distribution volume ratio relative to normal muscle and correlated to PARP-1 expression (P < 0.02, n = 10). Conclusion: This study of PARPi analog 18F-FTT showed that uptake kinetics invivo corresponded to expression of PARP-1 and that 18F-FTT quantitation is influenced by radiometabolites that are increasingly present late after injection. Radiometabolites can be controlled by using optimal image acquisition timing or normal muscle reference tissue modeling in dynamic imaging or a tumor-to-normal muscle ratio. Optimal image timing for tumor-to-normal muscle quantification in humans appears to be between 50 and 60 min after injection. Therefore, a clinically practical static imaging protocol commencing 45-55 min after injection may sufficiently balance 18F-FTT uptake with background clearance and radiometabolite interference for quantitative interpretation of PARP-1 expression invivo.

Predicting Malignancy in Adnexal Tumors With FAPI PET/CT and FDG PET/CT: A Case Report on a Borderline Ovarian Tumor.

A 54-year-old woman presented with a 55 × 64-mm tumor in the ovary with high [18F]FDG uptake on [18F]FDG PET/CT highly suggestive of ovarian cancer. Prior to surgery, the patient underwent [68Ga]-Ga- fibroblast activation protein inhibitor (FAPI)-46 PET/CT, which revealed low [68Ga]-Ga-FAPI-46 tumor uptake. Histopathology revealed a borderline ovarian tumor, which has low malignant potential and a 10-year survival rate greater than 93%. High [18F]FDG uptake is considered a fair predictor of malignancy in adnexal tumors. The present case demonstrates the potential superiority of [68Ga]-Ga-FAPI-46 PET/CT over [18F]-FDG PET/CT in differentiating malignant adnexal tumors from borderline ovarian tumors.

Novel [18F]FPG-interleukin-2 conjugate for monitoring immune checkpoint therapy with positron emission tomography

18F-interleukin-2 based PET imaging of activated T cells serves as a potential tool for non-invasive response prediction, treatment evaluation, and patient stratification in cancer immune checkpoint therapy. Herein, we report the radiolabelling of interleukin-2 (IL-2) with a novel arginine selective bioconjugation reagent, 4-[18F]fluorophenylglyoxal ([18F]FPG). Good non-decay corrected bioconjugation efficiencies of 29 ± 4 % (n = 5) were obtained for the [18F]FPG-IL-2. [18F]FPG-IL-2 uptake by the phytohemagglutinin-activated Jurkat cells (50.5 ± 1.2 %, n = 3) was significantly higher compared to the non-activated Jurkat cells (12.9 ± 1.1 %, n = 3). The [18F]FPG-IL-2 uptake was blocked by the pre-treatment of activated Jurkat cells with excess native IL-2 (22.3 ± 2.2 %, n = 3). Dynamic PET imaging and ex vivo biodistribution study of [18F]FPG-IL-2 in healthy and CT26 tumour bearing mice demonstrated hepatobiliary and renal clearance with minimal uptake in other organs and CT26 tumours. [18F]FPG-IL-2 PET imaging was applied to non-invasively monitor immune checkpoint therapy in CT26 tumour bearing mice, treated with IgG (control), ⍺PD-1 (monotherapy), and ⍺PD-1+⍺CTLA-4 (combination therapy). Significant uptake was observed in the spleens and tumours of the mice in the combination therapy group, which was associated with increased cytotoxic CD8+ T-cell infiltration and reduced tumour volumes. [18F]FPG-IL-2 based PET imaging has the potential to monitor immune checkpoint therapy.

Nuclear imaging of PD-L1 expression promotes the synergistic antitumor efficacy of targeted radionuclide therapy and immune checkpoint blockade.

In order to maximize synergistic effect of targeted radionuclide therapy (TRT) and immune checkpoint blockade (ICB) as well as reduce the toxicity, we pioneered a strategy guided by PD-L1-targeted nuclear medicine imaging for the combination of TRT and ICB towards precision cancer therapy. As a novel targeted radiotherapeutic agent, 177Lu-AB-3PRGD2 targeting integrin αvβ3 was developed to achieve sustained antitumor effect by introducing an albumin binder (AB) into the structure of 3PRGD2. The 177Lu-AB-3PRGD2 TRT as well as different types of combination therapies of 177Lu-AB-3PRGD2 TRT and anti-PD-L1 ICB were performed in animal models. The changes of PD-L1 expression in tumors after TRT were evaluated in vitro and in vivo by PD-L1-specific SPECT/CT imaging of 99mTc-MY1523. 177Lu-AB-3PRGD2 showed improved tumor uptake and prolonged tumor retention, leading to significantly enhanced tumor growth suppression. Moreover, 177Lu-AB-3PRGD2 TRT remodeled the tumor immune microenvironment by upregulating PD-L1 expression and increasing tumor-infiltrating CD8+ T cells, facilitating immunotherapy. We found that the anti-PD-L1 treatment was more effective during the upregulation of tumor PD-L1 expression, and the time window could be determined by 99mTc-MY1523 SPECT/CT. We developed a novel and long-acting radiotherapeutic agent 177Lu-AB-3PRGD2, and pioneered a strategy guided by PD-L1-targeted nuclear medicine imaging for the combination of TRT and ICB towards precision cancer therapy, optimizing the therapeutic efficacy and reducing the cost and potential toxicity risks. This strategy could also be adapted for clinical practice, combining conventional radiotherapy or chemotherapy with ICB to enhance therapeutic efficacy.

Rational modifications on N-(4-quinolinoyl)-Gly-2-cyanopyrrolidine to develop fibroblast activation protein-targeted radioligands with improved affinity and tumor uptake

Fibroblast activation protein (FAP) has been an attractive target for cancer imaging and therapy. Radiolabeled FAP-targeting ligands have shown promising results for clinical applications. However, further improvements are ongoing in pursuit of increasing tumor uptake, prolonging tumor residence, and maintenance good tumor–to–background contrast for extensive theranostic application. Achieving a higher affinity of the precursor is one of the ways in research. In this study, we designed a series of FAP inhibitors based on N-(4-quinolinoyl)-Gly-2-cyanopyrrolidine and found compound QI-18 with an IC50 value of 0.50 nM, which is a 6.5-fold increase in potency over that of UAMC-1110 (IC50 of 3.25 nM). QI-18 was then functionalized with a DOTA chelator to obtain the ligand CY03 for further radiolabeling with 68Ga to obtain the radiotracer [68Ga]Ga-CY03. In BALB/c nude mice bearing U87MG tumor models, [68Ga]Ga-CY03 exhibited a high and specific uptake (10.30 ± 0.63 % ID/g at 1 h post-injection and 9.28 ± 1.60 % ID/g at 2 h post-injection), which represented 3.2- and 4.1-fold increases over those for [68Ga]Ga-FAPI-04 (3.24 ± 0.53 % ID/g and 2.25 ± 0.33 % ID/g, respectively). [68Ga]Ga-CY03 also showed higher tumor–to–blood and tumor–to–kidney ratios (7.62 ± 0.44 and 2.59 ± 0.27, respectively) than [68Ga]Ga-FAPI-04 (2.38 ± 0.47 and 0.98 ± 0.19, respectively). The results indicate that [68Ga]Ga-CY03 is a promising imaging agent to target FAP.

64Cu]Cu-NOTA-Trastuzumab and [89Zr]Zr-DFO-Trastuzumab in Xenografts with Varied HER2 Expression.

Positron emission tomography (PET) has potential as a complementary technique to biomarker analysis, especially for human epidermal growth factor receptor 2 (HER2)-expressing tumors characterized by high heterogeneity. In this study, zirconium-89 (89Zr) and copper-64 (64Cu) labeled trastuzumab were employed to monitor varying levels of tumoral HER2 expression. Additionally, we studied the use of the cholesterol-depleting lovastatin as a pharmacological approach to enhance cell-surface HER2 expression in tumors with moderate to low HER2 levels, aiming to increase antibody accumulation in these tumor types. Both 89Zr- and 64Cu-labeled trastuzumab effectively monitor HER2 expression levels in xenografts exhibiting varying HER2 expression. No significant difference in tumor uptake was observed between 89Zr- or 64Cu-labeled trastuzumab, and tumor uptake for both radioimmunoconjugates positively correlated with HER2 protein levels. These findings underscore the potential of PET to monitor HER2 protein levels across heterogeneous tumors. Furthermore, our results suggest that further optimization of statin dosing and timing could offer a promising strategy to enhance trastuzumab accumulation in HER2-high, HER2-moderate, and HER2-low tumors.

Integrating molecular imaging and transcriptomic profiling in advanced HER2-positive breast cancer receiving trastuzumab emtansine (T-DM1): an analysis of the ZEPHIR clinical trial.

The ZEPHIR clinical trial evaluated the role of [89Zr]trastuzumab-PET/CT (HER2-PET/CT) and 2-[18F]fluoro-2-deoxy-D-glucose PET/CT ([18F]FDG-PET/CT) in predicting outcomes in patients with advanced HER2-positive breast cancer treated with trastuzumab emtansine (T-DM1). Here, we combined molecular/metabolic imaging and transcriptomic data to investigate the biological processes associated with [89Zr]trastuzumab and [18F]FDG uptake, and to dissect the mechanisms involved in T-DM1 resistance. RNA was extracted from metastasis biopsies obtained in the ZEPHIR trial. HER2-PET/CT and [18F]FDG-PET/CT imaging data of biopsied lesions were integrated with transcriptomic data. Lesions were compared based on the level of [89Zr]trastuzumab uptake as well as on the presence/absence of metabolic response, defined comparing baseline and on-treatment [18F]FDG-PET/CT. We analyzed matched transcriptomic and molecular/metabolic imaging data for 24 metastases. Genes and pathways involved in extracellular matrix (ECM) organization and glycosylphosphatidylinositol synthesis were enriched in lesions presenting low [89Zr]trastuzumab uptake. [18F]FDG uptake at baseline correlated with proliferation and immune-related processes. Hypoxia and ECM-related processes were enriched in lesions showing no metabolic response to T-DM1, while immune-related processes were associated with high [89Zr]trastuzumab uptake and metabolic response. Gene signatures including differentially expressed genes according to [89Zr]trastuzumab uptake and metabolic response showed predictive value in an external cohort. To our knowledge, this study represents the first correlative analysis between [89Zr]trastuzumab tumor uptake and gene expression profiling in humans. Our findings suggest a role of ECM in impairing [89Zr]trastuzumab tumor uptake and T-DM1 metabolic response in advanced HER2-positive breast cancer, highlighting the potential of molecular imaging to depict tumor microenvironment features.

CTR-FAPI PET enables precision management of medullary thyroid carcinoma.

Medullary thyroid carcinoma (MTC) can only be cured through the excision of all metastatic lesions, but 29-60% patients failed to localize the disease in the current clinical practice. Previously, we developed a fibroblast activation protein inhibitor (FAPI)-based covalent targeted radioligand (CTR) for improved detection sensitivity and accuracy. In this first-in-class clinical trial, we head-to-head compared [68Ga]Ga-CTR-FAPI PET-CT and [18F]FDG PET-CT in 50 MTC patients. The primary endpoint was the patient-based detection rate, with [68Ga]Ga-CTR-FAPI exhibiting higher detection than [18F]FDG (98% vs. 66%, p=0.0002). This improved detection was attributed to increased tumor uptake (SUVmax 11.71±9.16 vs. 2.55±1.73, p<0.0001). Diagnostic accuracy, validated on lesions with gold-standard pathology, was greater for [68Ga]Ga-CTR-FAPI compared to [18F]FDG (96.7% vs. 43.3%, p<0.0001). Notably, 32% patients altered management following [68Ga]Ga-CTR-FAPI PET-CT, and 66.7% patients changed their surgical plan. Overall, [68Ga]Ga-CTR-FAPI PET-CT provided superior detection and diagnostic accuracy compared to [18F]FDG PET-CT, enabling precision management of MTC patients.

52Mn-labelled Beta-cyclodextrin for Melanoma Imaging: A Proof-of-concept Preclinical Study.

As prostaglandin E2 (PGE2) and its receptors (EP2) are over-expressed on tumor cells and microenvironment, radiolabeled cyclodextrins targeting such biomolecules are valuable vector candidates in molecular cancer diagnostics. Using experimental melanoma models, we evaluated the in vivo imaging behavior of novel Manganese-52-labeled (52Mn) randomly methylated beta-cyclodextrin ([52Mn]Mn-DOTAGA-RAMEB) and compared it with the following well-established tumor-specific probes: melanocortin-1 receptor (MC1-R)-affine [68Ga]Ga-DOTA-NAPamide and PGE2 selective [68Ga]Ga-DOTAGA-RAMEB cyclodextrin. Post-injection of [68Ga]Ga-DOTA-NAPamide, [68Ga]Ga-DOTAGA-RAMEB, and [52Mn]Mn-DOTAGA-RAMEB into MC1-R positive B16F10 melanoma-bearing mice, tumor radio-pharmaceutical uptake was quantified in vivo and ex vivo using preclinical positron emission tomography (PET) and high-performance gamma counter. Although all tracers performed well in tumor identification, the highest standardized uptake values were detected in the [68Ga]Ga-DOTA-NAPamide scans. Corresponding to the ex vivo data, meaningful [52Mn]Mn-DOTAGA-RAMEB accumulation 1 h post-injection confirmed the tumor-targeting potential of the tracer. Temporal changes in PGE2/EP2 expression of the neoplasms may explain the significant differences observed between the tumor uptake of the two cyclodextrin probes and that of the 52Mn-labelled compound measured 1 h, 4 h, and 3 days post-injection (p≤0.01, p≤0.05). Although further pharmacokinetical optimization may be required, 52Mn-labelled cyclodextrin holds potential in melanoma diagnostics and the PET-based longitudinal assessment of tumor-associated PGE2/EP2 expression.

68Ga]Ga-FAPI04 Outperforms [18F]FDG PET/CT for Detecting Nodal Metastasis of Head and Neck Squamous Cell Carcinoma: A Single-Center Experience.

This study assesses fibroblast activated protein inhibitor (FAPI) targeted PET/CT imaging against [18F]FDG PET/CT (FDG PET) for detecting nodal involvement in head and neck squamous cell carcinoma (HNSCC), intending to improve diagnostic precision for metastatic lymph nodes and lay the groundwork for future investigations. Methods: Patients diagnosed with HNSCC were retrospectively enrolled. All patients underwent [68Ga]Ga-FAPI04 PET/CT (FAPI PET) and FDG PET within 6 d. Primary tumor, lymph nodes, and tracer uptake were visually and quantitatively compared. The metastatic lymph nodes were evaluated using patient-and lesion-based analyses, with biopsy or postoperative histopathological examination as the reference. Results: The cohort includes 24 patients (17 men, 7 women; mean age 60 ± 11.8 years) who underwent FDG and FAPI PET for preoperative diagnostic workup or restaging due to known recurrence of HNSCC. Lesions included 24 primary tumors, 54 cervical lymph nodes, and 5 metastases. Primary tumors exhibited significant uptake on both PET modalities (median maximum standardized uptake value [SUVmax]: FDG 19.4 ± 11.6, FAPI 16.9 ± 4.6), with no statistically significant difference (p > 0.5). For lymph nodes, FAPI and FDG PET showed median SUVmax of 9.18 ± 6.77 and 9.67 ± 6.5, respectively. The patient-based analysis found FDG PET sensitivity at 88.2% and FAPI PET at 94.1%, with FAPI PET specificity significantly higher (85.7% vs. 42.8% for FDG PET). Lesion-based analysis revealed FAPI PET sensitivity and specificity at 84.2% and 93.7%, respectively, contrasting FDG PET's at 81.5% and 25%, respectively. Conclusion: This study underscores the efficacy of FAPI PET in detecting primary tumors in HNSCC. Furthermore, FAPI PET shows improved specificity over FDG PET for metastatic lymph nodes advocating further investigations for integrating FAPI PET into HNSCC clinical protocols for its enhanced precision in detecting metastatic lymph nodes.

Synthesis and Evaluation of 68Ga- and 177Lu-Labeled [Pro14]bombesin(8-14) Derivatives for Detection and Radioligand Therapy of Gastrin-Releasing Peptide Receptor-Expressing Cancer.

The gastrin-releasing peptide receptor (GRPR) is overexpressed in a variety of cancers and represents a promising target for diagnosis and therapy. However, the extremely high accumulation in the pancreas observed for most of the clinically evaluated GRPR-targeted radiopharmaceuticals could limit their applications. In this study, we synthesized one GRPR antagonist (ProBOMB5) and two GRPR agonists (LW02056 and LW02057) by replacing the 4-thiazolidinecarboxylic acid (Thz14) residue in our previously reported GRPR-targeted tracers with Pro14. The 68Ga and 177Lu labeling were conducted in HEPES (2 M, pH 5.0) buffer and acetate (0.1 M, pH 4.5) buffer, respectively, and the radiolabeled products were obtained in a 24-57% decay-corrected radiochemical yield and >92% radiochemical purity. The binding affinities (Ki) of Ga-ProBOMB5, Ga-LW02056, Ga-LW02057, and Lu-ProBOMB5 were measured via in vitro competition binding assays and were 12.2 ± 1.89, 14.7 ± 4.81, 13.8 ± 2.24, and 13.6 ± 0.25 nM, respectively. The PET imaging and ex vivo biodistribution studies were conducted in PC-3 tumor-bearing mice at 1 h post injection. [68Ga]Ga-ProBOMB5, [68Ga]Ga-LW02056, and [68Ga]Ga-LW02057 enabled clear tumor visualization in PET images. The tumor uptake values of [68Ga]Ga-ProBOMB5, [68Ga]Ga-LW02056, and [68Ga]Ga-LW02057 were 12.4 ± 1.35, 8.93 ± 1.96, and 7.64 ± 0.55%ID/g, respectively, and their average pancreas uptake values were minimal (0.60-1.37%ID/g). Longitudinal SPECT imaging and ex vivo biodistribution studies were also conducted for [177Lu]Lu-ProBOMB5 and clinically validated [177Lu]Lu-RM2. Despite comparable tumor uptake at 1 h post injection ([177Lu]Lu-ProBOMB5:8.09 ± 1.70%ID/g; [177Lu]Lu-RM2:7.73 ± 0.96%ID/g), a faster clearance from PC-3 tumor xenografts was observed for [177Lu]Lu-ProBOMB5, leading to a lower radiation-absorbed dose delivered to tumors. Our data demonstrate that [68Ga]Ga-ProBOMB5 is a promising tracer for clinical translation for detecting GRPR-expressing tumor lesions. However, further optimizations are needed for [177Lu]Lu-ProBOMB5 to prolong tumor retention for therapeutic applications.

Head-to-head comparison of 68Ga-FAPI-46 PET/CT, 18F-FDG PET/CT, and contrast-enhanced CT for the detection of various tumors.

FAPI-PET/CT exhibits high tumor uptake and low background accumulation, enabling high-sensitivity tumor detection. We compared the diagnostic performance of 68Ga-FAPI-46 PET/CT plus contrast-enhanced CT (CE-CT), 18F-FDG PET/CT plus CE-CT, and standalone CE-CT in patients with various malignancies. 232 patients underwent 68Ga-FAPI-46 PET/CT,18F-FDG PET/CT, and CE-CT each within 4weeks. Detection rates were assessed by a blinded reader, with ≥ 2weeks between scans of the same patient to avoid recall bias. A sub-analysis of diagnostic performance was performed for 490 histopathologically validated lesions. Detection rates were compared using McNemar's test. Lesion-based detection rates in 68Ga-FAPI-46 PET/CT plus CE-CT, 18F-FDG PET/CT plus CE-CT, and CE-CT alone were 91.2% (1540/1688), 82.5% (1393/1688) and 60.2% (1016/1688). The detection rates were significantly higher for 68Ga-FAPI-46 PET/CT plus CE-CT than for 18F-FDG PET/CT plus CE-CT (p < 0.02 for primary lesions and p < 0.001 for total, abdominopelvic nodal, liver and other visceral lesions) and CE-CT (p < 0.0001 for total, primary, cervicothoracic nodal, abdominopelvic nodal, liver, other visceral, and bone lesions). In the sub-analysis, sensitivity, specificity, positive and negative predictive value, and accuracy were 61.3%, 96.7%, 81.4%, 91.4% and 90.0% for 68Ga-FAPI-46 PET/CT plus CE-CT, 57.0%, 95.7%, 75.7%, 90.5% and 88.4% for 18F-FDG PET/CT plus CE-CT, and 51.6%, 97.2%, 81.4%, 89.6% and 88.6% for CECT, respectively. 68Ga-FAPI-46 PET/CT plus CE-CT demonstrates a higher tumor detection rate than 18F-FDG PET/CT plus CE-CT and CE-CT in a diverse spectrum of malignancies, especially for primary, abdominopelvic nodal, liver, and other visceral lesions. Further studies on which entities draw particular benefit from 68Ga-FAPI-46 PET/CT are warranted to aid appropriate diagnostic workup. A total of N = 232 patients were analyzed. Of these, N = 50 patients were included in a prospective interventional trial (NCT05160051), and N = 175 in a prospective observational trial (NCT04571086) for correlation and clinical follow-up of PET findings; N = 7 patients were analyzed retrospectively.

PET Imaging Using 89Zr Labeled StarPEG Nanocarriers Reveals Heterogeneous Enhanced Permeability and Retention (EPR) in Prostate Cancer.

The enhanced permeability and retention (EPR) effect controls passive nanodrug uptake in tumors, and may provide a high tumor payload with prolonged retention for cancer treatment. However, EPR-mediated tumor uptake and distribution vary by cancer phenotype. Thus, we hypothesized that a companion PET-imaging surrogate may benefit EPR-mediated therapeutic drug delivery. We developed two 89Zr-radiolabeled nanocarriers based on 4-armed-starPEG40kDa with or without talazoparib (TLZ), a potent PARPi, as surrogates for the PEG-TLZ4 therapeutic scaffold. For PET imaging, PEG-DFB4 and PEG-DFB1-TLZ3 were radiolabeled with 89Zr by replacing one or all four TLZ on PEG-TLZ4 with deferoxamine B (DFB). The radiolabeled nanodrugs [89Zr]PEG-DFB4 and [89Zr]PEG-DFB1-TLZ3 were tested in vivo in prostate cancer subcutaneous xenografts (22Rv1, LTL-545, and LTL-610) and 22Rv1 metastatic models. Their EPR-mediated tumoral uptake and penetration was compared to CT26, a known EPR-high MicroPET/CT images, organ biodistribution, and calculated kinetic parameters showed high uptake in CT26 and LTL-545, moderate to low uptake in LTL-610 and 22Rv1. MicroPET/CT and high-resolution autoradiographic images showed nanocarrier penetration into highly permeable CT26, but heterogeneous peripheral accumulation was observed in LTL-545, LTL-610, and 22Rv1 subcutaneous xenografts and metastatic tumors. CD31 staining of tumor sections showed homogenous vascular development in CT26 tumors and heterogeneity in other xenografts. Both [89Zr]PEG-DFB4 and [89Zr]PEG-DFB1-TLZ3 showed similar accumulation and distribution in subcutaneous and metastatic tumor models. Both nanocarriers can measure tumor model passive uptake heterogeneity. Although heterogeneous, prostate cancer xenografts had low EPR. These starPEG nanocarriers could be used as PET imaging surrogates to predict drug delivery and efficacy.

Immuno-PET Imaging of EGFR with 64Cu-NOTA Panitumumab in Subcutaneous and Metastatic Nonsmall Cell Lung Cancer Xenografts.

Objective: About 65-90% of nonsmall cell lung cancer (NSCLC) express the epithelial growth factor receptor (EGFR) as a transmembrane protein that is activated by binding of specific ligands, including epidermal growth factor and transforming growth factor α (TGFα). Identifying EGFR as an oncogene has led to the development of anticancer therapeutics directed against EGFR, including the full-length human IgG2 monoclonal antibody panitumumab. The main goal of the present study was to investigate 64Cu-labeled panitumumab with immuno-PET in subcutaneous and metastatic EGFR-positive NSCLC xenografts. Methods: Bifunctional chelating agent 2-S-(4-isothiocyanatobenzyl)-1,4,7-triazacyclo-nonane-1,4,7-triacetic acid (NOTA-NCS) was attached to panitumumab. The number of chelators per panitumumab was determined using matrix-assisted laser desorption/ionization (MALDI) mass spectroscopy. The incorporation efficiency of 64Cu into NOTA-panitumumab was measured by using radio-TLC. EGFR-expressing epithelial-like H1299-luc+ NSCLC cells were used for in vitro and in vivo experiments. Cell uptake of [64Cu]Cu-NOTA-panitumumab was measured in the presence and absence of panitumumab. Subcutaneous and metastatic H1299-luc tumor models were grown in male NSG mice. The presence of tumors at lung and metastatic sites was analyzed by [18F]FLT PET. Immuno-PET with [64Cu]Cu-NOTA-panitumumab was performed as static PET imaging at 2, 24, and 48 h postinjection in both tumor models. Proof-of-target was confirmed by blocking experiments with panitumumab. Detailed ex vivo biodistribution experiments were performed in both animal tumor models to confirm biodistribution profiles obtained by immuno-PET imaging. Results: MALDI analysis confirmed the attachment of ∼1.5 NOTA per antibody. Radiolabeling efficiency with [64Cu]CuCl2 was 93.8 ± 5.7% and a molar activity of 0.65 MBq/μg. Cellular uptake studies with [64Cu]Cu-NOTA-panitumumab in H1299 cells demonstrated increasing uptake over time, reaching 29.1 ± 2.9% radioactivity(Bq)/mg protein (n = 3) and plateauing at 45 min. Addition of 25 μg of panitumumab reduced radioligand uptake to 1.22 ± 0.06% radioactivity/mg protein (n = 3). PET imaging revealed high uptake of [64Cu]Cu-NOTA-panitumumab in subcutaneous tumors: Standardized uptake values (SUV)mean reached 4.70 ± 0.42 and 5.37 ± 0.40 (n = 5) after 24 and 48 h postinjection, respectively. Administration of 1 mg panitumumab reduced tumor uptake significantly to 1.94 ± 0.22 and 1.66 ± 0.08 (n = 4; p < 0.001). In the metastatic model, the following SUVmean were analyzed from liver and lung lesions: 5.55 ± 0.34 and 6.28 ± 0.46 (both n = 23 lesions from 6 mice) after 24 and 48 h postinjection, which was also significantly reduced to 2.53 ± 0.39 and 2.31 ± 0.15 (both n = 16 lesions from 4 mice; p < 0.001) after injection of 1 mg panitumumab. Detailed ex vivo biodistribution confirmed immuno-PET analysis in both models. Panitumumab reduced radioactivity uptake into subcutaneous tumors from 11.01 ± 0.72 (n = 4) to 3.67 ± 0.33% ID/g (n = 5; p < 0.001), and in metastatic liver lesions from 29.44 ± 8.14 (n = 4) to 8.35 ± 1.30% ID/g (n = 5; p < 0.001), respectively. Conclusions: [64Cu]Cu-NOTA-panitumumab was successfully used for immuno-PET imaging of EGFR-expressing subcutaneous and metastatic NSCLC tumors. This result represents the basis for developing radiotheranostics for targeting EGFR in cancers and for selecting the right patients for the right treatment at the right time.

ESOMA-DM1, a Maytansinoid-Based Theranostic Small-Molecule Drug Conjugate for Neuroendocrine Tumors.

Background: The main challenges of conventional chemotherapy lie in its lack of selectivity and specificity, leading to significant side effects. Using a small-molecule drug conjugate (SMDC) ensures specific delivery of a cytotoxic drug to the tumor site by coupling it to a targeting vector. This promising strategy can be applied to neuroendocrine tumors (NETs) by choosing a targeting vector that binds specifically to somatostatin receptor subtype 2 (SSTR2). Additionally, incorporation of a bifunctional chelate into the molecule enables complexation of both diagnostic and therapeutic radionuclides. Thus, it facilitates monitoring of the distribution of the SMDC in the body and allows for the implementation of combination therapy. In our study, we designed eSOMA-DM1, a SMDC combining the SSTR2-targeted octreotate peptide and the cytotoxic agent DM1 via a chelate-bridged linker (N3-Py-DOTAGA). This approach warrants conjugation of the targeting vector and the drug at opposite sites to avoid undesired steric hindrance effects. Methods: Synthesis of the DM1 moiety (4) involved a three-step synthetic route, followed by the conjugation to the cyclic peptide, N3-Py-DOTAGA-d-Phe-cyclo[Cys-Tyr-d-Trp-Lys-Thr-Cys]-Thr-OH, through a copper-free click reaction, resulting in eSOMA-DM1. Subsequent labeling with [111In]InCl3 gave a high radiochemical yield and purity. In vitro assessments of eSOMA-DM1 binding, uptake, and internalization were conducted in SSTR2-transfected U2OS cells. Ex vivo biodistribution and fluorescence imaging were performed in H69-tumor bearing mice. Results: eSOMA-DM1 exhibited an IC50 value for SSTR2 similar to the gold standard DOTA-TATE. The uptake of [111In]In-eSOMA-DM1 in U2OS.SSTR2 cells was 1.2-fold lower than that of [111In]In-DOTA-TATE. Tumor uptake in H69-xenografted mice was higher for [111In]In-eSOMA-DM1 at all-time points compared to [111In]In-DOTA-TATE. Prolonged blood circulation led to increased accumulation of [111In]In-eSOMA-DM1 in highly vascularized tissues, such as the lungs, skin, and heart. Excretion through the kidneys, liver, and spleen was also observed. Conclusion: eSOMA-DM1 is a SMDC developed for NET showing promising characteristics in vitro. However, the in vivo results obtained with [111In]In-eSOMA-DM1 suggest the need for adjustments to optimize its distribution.