Search Results (19)

Prostate-specific membrane antigen (PSMA)-based low-molecular-weight agents using beta(β)-particle-emitting radiopharmaceuticals is a new treatment paradigm for patients with metastatic castration-resistant prostate cancer. Although results have been encouraging, there is a need to improve the tumor residence time of current PSMA-based radiotherapeutics. Albumin-binding moieties have been used strategically to enhance the tumor uptake and retention of existing PSMA-based investigational agents. Previously, we developed a series of PSMA-based, β-particle-emitting, low-molecular-weight compounds. From this series, ¹⁷⁷Lu-L1 was selected as the lead agent because of its reduced off-target radiotoxicity in preclinical studies. The ligand L1 contains a PSMA-targeting Lys-Glu urea moiety with an N-bromobenzyl substituent in the ε-amino group of Lys. Here, we structurally modified ¹⁷⁷Lu-L1 to improve tumor targeting using two known albumin-binding moieties, 4-(p-iodophenyl) butyric acid moiety (IPBA) and ibuprofen (IBU), and evaluated the effects of linker length and composition. Six structurally related PSMA-targeting ligands (Alb-L1–Alb-L6) were synthesized based on the structure of ¹⁷⁷Lu-L1. The ligands were assessed for in vitro binding affinity and were radiolabeled with ¹⁷⁷Lu following standard protocols. All ¹⁷⁷Lu-labeled analogs were studied in cell uptake and selected cell efficacy studies. In vivo pharmacokinetics were investigated by conducting tissue biodistribution studies for ¹⁷⁷Lu-Alb-L2–¹⁷⁷Lu-Alb-L6 (2 h, 24 h, 72 h, and 192 h) in male NSG mice bearing human PSMA+ PC3 PIP and PSMA− PC3 flu xenografts. Preliminary therapeutic ratios of the agents were estimated from the area under the curve (AUC_0-192h) of the tumors, blood, and kidney uptake values. Compounds were obtained in >98% radiochemical yields and >99% purity. PSMA inhibition constants (K_is) of the ligands were in the ≤10 nM range. The long-linker-based agents, ¹⁷⁷Lu-Alb-L4 and ¹⁷⁷Lu-Alb-L5, displayed significantly higher tumor uptake and retention (p < 0.001) than the short-linker-bearing ¹⁷⁷Lu-Alb-L2 and ¹⁷⁷Lu-Alb-L3 and a long polyethylene glycol (PEG) linker-bearing agent, ¹⁷⁷Lu-Alb-L6. The area under the curve (AUC_0-192h) of the PSMA+ PC3 PIP tumor uptake of ¹⁷⁷Lu-Alb-L4 and ¹⁷⁷Lu-Alb-L5 were >4-fold higher than ¹⁷⁷Lu-Alb-L2, ¹⁷⁷Lu-Alb-L3, and ¹⁷⁷Lu-Alb-L6, respectively. Also, the PSMA+ PIP tumor uptake (AUC_0-192h) of ¹⁷⁷Lu-Alb-L2 and ¹⁷⁷Lu-Alb-L3 was ~1.5-fold higher than ¹⁷⁷Lu-Alb-L6. However, the lowest blood AUC_0-192h and kidney AUC_0-192h were associated with ¹⁷⁷Lu-Alb-L6 from the series. Consequently, ¹⁷⁷Lu-Alb-L6 displayed the highest ratios of AUC(tumor)-to-AUC(blood) and AUC(tumor)-to-AUC(kidney) values from the series. Among the other agents, ¹⁷⁷Lu-Alb-L4 demonstrated a nearly similar ratio of AUC(tumor)-to-AUC(blood) as ¹⁷⁷Lu-Alb-L6. The tumor-to-blood ratio was the dose-limiting therapeutic ratio for all of the compounds. Conclusions: ¹⁷⁷Lu-Alb-L4 and ¹⁷⁷Lu-Alb-L6 showed high tumor uptake in PSMA+ tumors and tumor-to-blood ratios. The data suggest that linker length and composition can be modulated to generate an optimized therapeutic agent. Full article

[¹⁸F]DCFPyL is increasingly used for prostate-specific membrane antigen (PSMA) mediated imaging of men with biochemically recurrent prostate cancer (BRPCa). In this meta-analysis, which is updated with the addition of multiple new studies, including the definitive phase III CONDOR trial, we discuss the detection efficiency of [¹⁸F]DCFPyL in BRPCa patients. PubMed was searched on 29 September 2022. Studies evaluating the diagnostic performance of [¹⁸F]DCFPyL among patients with BRPCa were included. The overall pooled detection rate with a 95% confidence interval (95% CI) was calculated among all included studies and stratified among patients with PSA ≥ 2 vs. <2 ng/mL and with PSA ≥ 0.5 vs. <0.5 ng/mL. The association of detection efficiency with pooled PSA doubling time from two studies was calculated. Seventeen manuscripts, including 2252 patients, met the inclusion criteria and were used for data extraction. A previous meta-analysis reported that the pooled detection rate was 0.81 (95% CI: 0.77–0.85), while our study showed a pooled overall detection rate of 0.73 (95% CI: 0.66–0.79). An increased proportion of positive scans were found in patients with PSA ≥ 2 vs. <2 ng/mL and PSA ≥ 0.5 vs. <0.5 ng/mL. No significant difference was found in detection efficiency between those with PSA doubling time ≥ 12 vs. <12 months. Detection efficiency is statistically related to serum PSA levels but not to PSA doubling time based on available data. The detection efficiency of [¹⁸F]DCFPyL in men with BRPCa has trended down since a previous meta-analysis, which may reflect increasingly stringent inclusion criteria for studies over time. Full article

The use of prostate-specific membrane antigen targeted PET imaging for the evaluation of prostate cancer has increased significantly in the last couple of decades. When evaluating these imaging findings based on the PSMA reporting and data system version 1.0, which categorize lesions based on their likelihood of prostate cancer involvement, PSMA-RADS-3A lesions are commonly seen, which are indeterminate for the presence of disease. A total of 28 patients with 171 PSMA-RADS-3A lesions on ¹⁸F-DCFPyL PET/CT scans from June 2016 to May 2017 who had follow-up cross-sectional imaging over time were included in this study. The PSA levels of patients with PSMA-RADS-3A lesions were categorized into four groups, 0–0.2, 0.2–1, 1–2, and >2 ng/mL. The pre-operative Gleason score of these patients was categorized into two groups, Gleason score < 7 or ≥7. The median age for these patients was 72.5 years (range 59–81). The median PSA value for patients with positive lesions was significantly higher than those with negative lesions (5.8 ng/mL vs. 0.2 ng/mL, p < 0.0001). The lesion positivity rate was significantly higher in patients with PSA > 1 ng/mL (18.2% vs. 81.9%, p < 0.001). On ROC analysis, the highest classification accuracy was seen at PSA ≥ 0.6 ng/mL of 80.12% (95% CI = 73.69–86.16%), and the area under the curve was 71.32% (95% CI = 61.9–80.7%, p < 0.0001). A total of 96.4% (108/112) of patients with positive lesions and 86.4% (51/59) of patients with negative lesions had a PSMA-RADS-4/5 lymph node on the initial ¹⁸F-DCFPyL PET/CT scan (p = 0.02). In patients with a Gleason score ≥ 7, the presence of positive PSMA-RADS-3A lesions was higher, compared to negative PSMA-RADS-3A lesions (p = 0.049). Higher PSA levels in patients with PSMA-RADS-3A lesions can point towards the presence of true positivity. PSA levels may be considered in deciding whether to call an indeterminate lesion on PSMA PET. Full article

While machine learning (ML) methods may significantly improve image quality for SPECT imaging for the diagnosis and monitoring of Parkinson’s disease (PD), they require a large amount of data for training. It is often difficult to collect a large population of patient data to support the ML research, and the ground truth of lesion is also unknown. This paper leverages a generative adversarial network (GAN) to generate digital brain phantoms for training ML-based PD SPECT algorithms. A total of 594 PET 3D brain models from 155 patients (113 male and 42 female) were reviewed and 1597 2D slices containing the full or a portion of the striatum were selected. Corresponding attenuation maps were also generated based on these images. The data were then used to develop a GAN for generating 2D brain phantoms, where each phantom consisted of a radioactivity image and the corresponding attenuation map. Statistical methods including histogram, Fréchet distance, and structural similarity were used to evaluate the generator based on 10,000 generated phantoms. When the generated phantoms and training dataset were both passed to the discriminator, similar normal distributions were obtained, which indicated the discriminator was unable to distinguish the generated phantoms from the training datasets. The generated digital phantoms can be used for 2D SPECT simulation and serve as the ground truth to develop ML-based reconstruction algorithms. The cumulated experience from this work also laid the foundation for building a 3D GAN for the same application. Full article

The chemokine receptor 4 (CXCR4) is a promising diagnostic and therapeutic target for the management of various cancers. CXCR4 has been utilized in immunotherapy, targeted drug delivery, and endoradiotherapy. Poly(amidoamine) [PAMAM] dendrimers are well-defined polymers with unique properties that have been used in the fabrication of nanomaterials for several biomedical applications. Here, we describe the formulation and pharmacokinetics of generation-5 CXCR4-targeted PAMAM (G5-X4) dendrimers. G5-X4 demonstrated an IC₅₀ of 0.95 nM to CXCR4 against CXCL12-Red in CHO-SNAP-CXCR4 cells. Single-photon computed tomography/computed tomography imaging and biodistribution studies of ¹¹¹In-labeled G5-X4 showed enhanced uptake in subcutaneous U87 glioblastoma tumors stably expressing CXCR4 with 8.2 ± 2.1, 8.4 ± 0.5, 11.5 ± 0.9, 10.4 ± 2.6, and 8.8 ± 0.5% injected dose per gram of tissue at 1, 3, 24, 48, and 120 h after injection, respectively. Specific accumulation of [¹¹¹In]G5-X4 in CXCR4-positive tumors was inhibited by the peptidomimetic CXCR4 inhibitor, POL3026. Our results demonstrate that while CXCR4 targeting is beneficial for tumor accumulation at early time points, differences in tumor uptake are diminished over time as passive accumulation takes place. This study further confirms the applicability of PAMAM dendrimers for imaging and therapeutic applications. It also emphasizes careful consideration of image acquisition and/or treatment times when designing dendritic nanoplatforms for tumor targeting. Full article

We have synthesized a series of 10 new, PSMA-targeted, near-infrared imaging agents intended for use in vivo for fluorescence-guided surgery (FGS). Compounds were synthesized from the commercially available amine-reactive active NHS ester of DyLight800. We altered the linker between the PSMA-targeting urea moiety and the fluorophore with a view to improve the pharmacokinetics. Chemical yields for the conjugates ranged from 51% to 86%. The K_i values ranged from 0.10 to 2.19 nM. Inclusion of an N-bromobenzyl substituent at the ε-amino group of lysine enhanced PSMA⁺ PIP tumor uptake, as did hydrophilic substituents within the linker. The presence of a polyethylene glycol chain within the linker markedly decreased renal uptake. In particular, DyLight800-10 demonstrated high specific uptake relative to background signal within kidney, confirmed by immunohistochemistry. These compounds may be useful for FGS in prostate, renal or other PSMA-expressing cancers. Full article

The inherent instability of nucleic acids within serum and the tumor microenvironment necessitates a suitable vehicle for non-viral gene delivery to malignant lesions. A specificity-conferring mechanism is also often needed to mitigate off-target toxicity. In the present study, we report a stable and efficient redox-sensitive nanoparticle system with a unique core–shell structure as a DNA carrier for cancer theranostics. Thiolated polyethylenimine (PEI-SH) is complexed with DNA through electrostatic interactions to form the core, and glycol chitosan-modified with succinimidyl 3-(2-pyridyldithio)propionate (GCS-PDP) is grafted on the surface through a thiolate-disulfide interchange reaction to form the shell. The resulting nanoparticles, GCS-PDP/PEI-SH/DNA nanoparticles (GNPs), exhibit high colloid stability in a simulated physiological environment and redox-responsive DNA release. GNPs not only show a high and redox-responsive cellular uptake, high transfection efficiency, and low cytotoxicity in vitro, but also exhibit selective tumor targeting, with minimal toxicity, in vivo, upon systemic administration. Such a performance positions GNPs as viable candidates for molecular-genetic imaging and theranostic applications. Full article

(1) Background: Prostate-specific membrane antigen (PSMA)-directed radioligand therapy (RLT) has shown remarkable results in patients with advanced prostate cancer. We aimed to evaluate the toxicity profile of the PSMA ligand [¹⁷⁷Lu]Lu-PSMA I&T. (2) Methods: 49 patients with metastatic, castration-resistant prostate cancer treated with at least three cycles of [¹⁷⁷Lu]Lu-PSMA I&T were evaluated. Prior to and after RLT, we compared leukocytes, hemoglobin, platelet counts, and renal functional parameters (creatinine, eGFR, n = 49; [^99mTc]-MAG3-derived tubular extraction rate (TER), n = 42). Adverse events were classified according to the Common Terminology Criteria for Adverse Events (CTCAE) v5.0 and KDIGO Society. To identify predictive factors, we used Spearman’s rank correlation coefficient. (3) Results: A substantial fraction of the patients already showed impaired renal function and reduced leukocyte counts at baseline. Under RLT, 11/49 (22%) patients presented with nephrotoxicity CTCAE I or II according to creatinine, but 33/49 (67%) according to eGFR. Only 5/42 (13%) showed reduced TER, defined as <70% of the age-adjusted mean normal values. Of all renal functional parameters, absolute changes of only 2% were recorded. CTCAE-based re-categorization was infrequent, with creatinine worsening from I to II in 2/49 (4.1%; GFR, 1/49 (2%)). Similar results were recorded for KDIGO (G2 to G3a, 1/49 (2%); G3a to G3b, 2/49 (4.1%)). After three cycles, follow-up eGFR correlated negatively with age (r = −0.40, p = 0.005) and the eGFR change with Gleason score (r = −0.35, p < 0.05) at baseline. Leukocytopenia CTCAE II occurred only in 1/49 (2%) (CTCAE I, 20/49 (41%)) and CTCAE I thrombocytopenia in 7/49 (14%), with an absolute decrease of 15.2% and 16.6% for leukocyte and platelet counts. Anemia CTCAE II occurred in 10/49 (20%) (CTCAE I, 36/49 (73%)) with a decrease in hemoglobin of 4.7%. (4) Conclusions: After PSMA-targeted therapy using [¹⁷⁷Lu]Lu-PSMA I&T, no severe (CTCAE III/IV) toxicities occurred, thereby demonstrating that serious adverse renal or hematological events are unlikely to be a frequent phenomenon with this agent. Full article

Prostate-specific membrane antigen (PSMA)-directed positron emission tomography/computed tomography (PET/CT) is increasingly utilized for staging of men with prostate cancer (PC). To increase interpretive certainty, the standardized PSMA reporting and data system (RADS) has been proposed. Using PSMA-RADS, we characterized lesions in 18 patients imaged with ¹⁸F-PSMA-1007 PET/CT for primary staging and determined the stability of semi-quantitative parameters. Six hundred twenty-three lesions were categorized according to PSMA-RADS and manually segmented. In this context, PSMA-RADS-3A (soft-tissue) or -3B (bone) lesions are defined as being indeterminate for the presence of PC. For PMSA-RADS-4 and -5 lesions; however, PC is highly likely or almost certainly present [with further distinction based on absence (PSMA-RADS-4) or presence (PSMA-RADS-5) of correlative findings on CT]. Standardized uptake values (SUV_max, SUV_peak, SUV_mean) were recorded, and volumetric parameters [PSMA-derived tumor volume (PSMA-TV); total lesion PSMA (TL-PSMA)] were determined using different maximum intensity thresholds (MIT) (40 vs. 45 vs. 50%). SUV_max was significantly higher in PSMA-RADS-5 lesions compared to all other PSMA-RADS categories (p ≤ 0.0322). In particular, the clinically challenging PSMA-RADS-3A lesions showed significantly lower SUV_max and SUV_peak compared to the entire PSMA-RADS-4 or -5 cohort (p < 0.0001), while for PSMA-RADS-3B this only applies when compared to the entire PSMA-RADS-5 cohort (p < 0.0001), but not to the PSMA-RADS-4 cohort (SUV_max, p = 0.07; SUV_peak, p = 0.08). SUV_mean (p = 0.30) and TL-PSMA (p = 0.16) in PSMA-RADS-5 lesions were not influenced by changing the MIT, while PSMA-TV showed significant differences when comparing 40 vs. 50% MIT (p = 0.0066), which was driven by lymph nodes (p = 0.0239), but not bone lesions (p = 0.15). SUV_max was significantly higher in PSMA-RADS-5 lesions compared to all other PSMA-RADS categories in ¹⁸F-PSMA-1007 PET/CT. As such, the latter parameter may assist the interpreting molecular imaging specialist in assigning the correct PSMA-RADS score to sites of disease, thereby increasing diagnostic certainty. In addition, changes of the MIT in PSMA-RADS-5 lesions had no significant impact on SUV_mean and TL-PSMA in contrast to PSMA-TV. Full article

The 2-deoxy-d-[¹⁸F]fluoro-D-glucose (FDG) positron emission tomography/computed tomography (PET/CT) is widely utilized to assess the vascular and articular inflammatory burden of patients with a suspected diagnosis of rheumatic disease. We aimed to elucidate the impact of [¹⁸F]FDG PET/CT on change in initially suspected diagnosis in patients at the time of the scan. Thirty-four patients, who had undergone [¹⁸F]FDG PET/CT, were enrolled and the initially suspected diagnosis prior to [¹⁸F]FDG PET/CT was compared to the final diagnosis. In addition, a semi-quantitative analysis including vessel wall-to-liver (VLR) and joint-to-liver (JLR) ratios was also conducted. Prior to [¹⁸F]FDG PET/CT, 22/34 (64.7%) of patients did not have an established diagnosis, whereas in 7/34 (20.6%), polymyalgia rheumatica (PMR) was suspected, and in 5/34 (14.7%), giant cell arteritis (GCA) was suspected by the referring rheumatologists. After [¹⁸F]FDG PET/CT, the diagnosis was GCA in 19/34 (55.9%), combined GCA and PMR (GCA + PMR) in 9/34 (26.5%) and PMR in the remaining 6/34 (17.6%). As such, [¹⁸F]FDG PET/CT altered suspected diagnosis in 28/34 (82.4%), including in all unclear cases. VLR of patients whose final diagnosis was GCA tended to be significantly higher when compared to VLR in PMR (GCA, 1.01 ± 0.08 (95%CI, 0.95–1.1) vs. PMR, 0.92 ± 0.1 (95%CI, 0.85–0.99), p = 0.07), but not when compared to PMR + GCA (1.04 ± 0.14 (95%CI, 0.95–1.13), p = 1). JLR of individuals finally diagnosed with PMR (0.94 ± 0.16, (95%CI, 0.83–1.06)), however, was significantly increased relative to JLR in GCA (0.58 ± 0.04 (95%CI, 0.55–0.61)) and GCA + PMR (0.64 ± 0.09 (95%CI, 0.57–0.71); p < 0.0001, respectively). In individuals with a suspected diagnosis of rheumatic disease, an inflammatory-directed [¹⁸F]FDG PET/CT can alter diagnosis in the majority of the cases, particularly in subjects who were referred because of diagnostic uncertainty. Semi-quantitative assessment may be helpful in establishing a final diagnosis of PMR, supporting the notion that a quantitative whole-body read-out may be useful in unclear cases. Full article

Tracing its roots back to the 1940s, theranostics in nuclear oncology has proved successful mainly due to the beneficial effects of image-guided therapeutic concepts for patients afflicted with a variety of different cancers. The majority of these treatments are not only characterized by substantial prolongation of progression-free and overall survival, but are also generally safe, rendering theranostic agents as an attractive treatment option in various clinical scenarios in oncology. In this Special Issue Novel Theranostic Agents, nine original articles from around the globe provide further evidence on the use of the theranostic concept for neuroendocrine neoplasm (NEN), prostate cancer (PC), meningioma, and neuroblastoma. The investigated diagnostic and therapeutic radiotracers target not only established structures, such as somatostatin receptor, prostate-specific membrane antigen or norepinephrine transporter, but also recently emerging targets such as the C-X-C motif chemokine receptor 4. Moreover, the presented original articles also combine the concept of theranostics with in-depth read-out techniques such as radiomics or novel reconstruction algorithms on pretherapeutic scans, e.g., for outcome prediction. Even 80 years after its initial clinical introduction, theranostics in oncology continues to thrive, now more than ever. Full article

Purpose: Prostate-specific membrane antigen (PSMA) positron emission tomography (PET) is emerging as an important modality for imaging patients with prostate cancer (PCa). As with any imaging modality, indeterminate findings will arise. The PSMA reporting and data system (PSMA-RADS) version 1.0 codifies indeterminate soft tissue findings with the PSMA-RADS-3A moniker. We investigated the role of point-spread function (PSF) reconstructions on categorization of PSMA-RADS-3A lesions. Methods: This was a post hoc analysis of an institutional review board approved prospective trial. Around 60 min after the administration of 333 MBq (9 mCi) of PSMA-targeted ¹⁸F-DCFPyL, patients underwent PET/computed tomography (CT) acquisitions from the mid-thighs to the skull vertex. The PET data were reconstructed with and without PSF. Scans were categorized according to PSMA-RADS version 1.0, and all PSMA-RADS-3A lesions on non-PSF images were re-evaluated to determine if any could be re-categorized as PSMA-RADS-4. The maximum standardized uptake values (SUVs) of the lesions, mean SUVs of blood pool, and the ratios of those values were determined. Results: A total of 171 PSMA-RADS-3A lesions were identified in 30 patients for whom both PSF reconstructions and cross-sectional imaging follow-up were available. A total of 13/171 (7.6%) were re-categorized as PSMA-RADS-4 lesions with PSF reconstructions. A total of 112/171 (65.5%) were found on follow-up to be true positive for PCa, with all 13 of the re-categorized lesions being true positive on follow-up. The lesions that were re-categorized trended towards having higher SUV_max-lesion and SUV_max-lesion/SUV_mean-blood-pool metrics, although these relationships were not statistically significant. Conclusions: The use of PSF reconstructions for ¹⁸F-DCFPyL PET can allow the appropriate re-categorization of a small number of indeterminate PSMA-RADS-3A soft tissue lesions as more definitive PSMA-RADS-4 lesions. The routine use of PSF reconstructions for PSMA-targeted PET may be of value at those sites that utilize this technology. Full article

We aimed to elucidate the diagnostic potential of the C-X-C motif chemokine receptor 4 (CXCR4)-directed positron emission tomography (PET) tracer ⁶⁸Ga-Pentixafor in patients with poorly differentiated neuroendocrine carcinomas (NEC), relative to the established reference standard ¹⁸F-FDG PET/computed tomography (CT). In our database, we retrospectively identified 11 treatment-naïve patients with histologically proven NEC, who underwent ¹⁸F-FDG and CXCR4-directed PET/CT for staging and therapy planning. The images were analyzed on a per-patient and per-lesion basis and compared to immunohistochemical staining (IHC) of CXCR4 from PET-guided biopsies. ⁶⁸Ga-Pentixafor visualized tumor lesions in 10/11 subjects, while¹⁸F-FDG revealed sites of disease in all 11 patients. Although weak to moderate CXCR4 expression could be corroborated by IHC in 10/11 cases, ¹⁸F-FDG PET/CT detected significantly more tumor lesions (102 vs. 42; total lesions, n = 107; p < 0.001). Semi-quantitative analysis revealed markedly higher ¹⁸F-FDG uptake as compared to ⁶⁸Ga-Pentixafor (maximum and mean standardized uptake values (SUV) and tumor-to-background ratios (TBR) of cancerous lesions, SUV_max: 12.8 ± 9.8 vs. 5.2 ± 3.7; SUV_mean: 7.4 ± 5.4 vs. 3.1 ± 3.2, p < 0.001; and, TBR 7.2 ± 7.9 vs. 3.4 ± 3.0, p < 0.001). Non-invasive imaging of CXCR4 expression in NEC is inferior to the reference standard ¹⁸F-FDG PET/CT. Full article

Radiotherapy is a common approach for the treatment of a wide variety of cancer types. Available data indicate that nanoparticles can enhance the effect of radiotherapy. We report the use of human mesenchymal stem cells to selectively deliver gold nanoparticles (GNPs) to MDA-MB-231 breast tumor xenografts in mice for the purpose of enhancing the effect of radiation therapy. Targeted delivery of GNPs to the tumor site, followed by irradiation of the tumor, enabled control of tumor growth. The results indicate that tumor-selective GNP delivery by human mesenchymal stem cells may represent a viable way to enhance the effectiveness of radiotherapy. Full article

Prostate-Specific Membrane Antigen (PSMA) is an established biomarker for the imaging and experimental therapy of prostate cancer (PCa), as it is strongly upregulated in high-grade primary, androgen-independent, and metastatic lesions. Here, we report on the development and functional characterization of recombinant single-chain Fv (scFv) and Fab fragments derived from the 5D3 PSMA-specific monoclonal antibody (mAb). These fragments were engineered, heterologously expressed in insect S2 cells, and purified to homogeneity with yields up to 20 mg/L. In vitro assays including ELISA, immunofluorescence and flow cytometry, revealed that the fragments retain the nanomolar affinity and single target specificity of the parent 5D3 antibody. Importantly, using a murine xenograft model of PCa, we verified the suitability of fluorescently labeled fragments for in vivo imaging of PSMA-positive tumors and compared their pharmacokinetics and tissue distribution to the parent mAb. Collectively, our data provide an experimental basis for the further development of 5D3 recombinant fragments for future clinical use. Full article