Systematic Review of Cancer Targeting by Nanoparticles Revealed a Global Association between Accumulation in Tumors and Spleen

Active targeting of nanoparticles toward tumors is one of the most rapidly developing topics in nanomedicine. Typically, this strategy involves the addition of cancer-targeting biomolecules to nanoparticles, and studies on this topic have mainly focused on the localization of such formulations in tumors. Here, the analysis of the factors determining efficient nanoparticle targeting and therapy, various parameters such as types of targeting molecules, nanoparticle type, size, zeta potential, dose, and the circulation time are given. In addition, the important aspects such as how active targeting of nanoparticles alters biodistribution and how non-specific organ uptake influences tumor accumulation of the targeted nanoformulations are discussed. The analysis reveals that an increase in tumor accumulation of targeted nanoparticles is accompanied by a decrease in their uptake by the spleen. There is no association between targeting-induced changes of nanoparticle concentrations in tumors and other organs. The correlation between uptake in tumors and depletion in the spleen is significant for mice with intact immune systems in contrast to nude mice. Noticeably, modulation of splenic and tumor accumulation depends on the targeting molecules and nanoparticle type. The median survival increases with the targeting-induced nanoparticle accumulation in tumors; moreover, combinatorial targeting of nanoparticle drugs demonstrates higher treatment efficiencies. Results of the comprehensive analysis show optimal strategies to enhance the efficiency of actively targeted nanoparticle-based medicines.

Spike protein multiorgan tropism suppressed by antibodies targeting SARS-CoV-2

While there is SARS-CoV-2 multiorgan tropism in severely infected COVID-19 patients, it’s unclear if this occurs in healthy young individuals. In addition, for antibodies that target the spike protein (SP), it’s unclear if these reduce SARS-CoV-2/SP multiorgan tropism equally. We used fluorescently labeled SP-NIRF to study viral behavior, using an in vivo dynamic imaging system and ex in vivo tissue analysis, in young mice. We found a SP body-wide biodistribution followed by a slow regional elimination, except for the liver, which showed an accumulation. SP uptake was highest for the lungs, and this was followed by kidney, heart and liver, but, unlike the choroid plexus, it was not detected in the brain parenchyma or CSF. Thus, the brain vascular barriers were effective in restricting the entry of SP into brain parenchyma in young healthy mice. While both anti-ACE2 and anti-SP antibodies suppressed SP biodistribution and organ uptake, anti-SP antibody was more effective. By extension, our data support the efficacy of these antibodies on SARS-CoV-2 multiorgan tropism, which could determine COVID-19 organ-specific outcomes.

Development and preclinical evaluation of novel fluorinated ammonium salts for PET myocardial perfusion imaging

We previously presented the radiolabeled ammonium salt [11C]-dimethyl diphenylammonium trifluoromethanesulfonate ([11C]DMDPA) as a potential novel PET-MPI agent. The current study aimed to increase the clinical applicability of PET-MPI by designing and synthesizing fluorinated ammonium salt derivatives. Four fluorinated DMDPA derivatives and two quinolinium salt analogs were radiolabeled. The dynamic distribution in vivo, following injection of each derivative into male SD rats, was evaluated using small-animal dedicated PET/CT. Organ uptake after injection of [18F]fluoroethylquinolinium acetate ([18F]FEtQ) was examined ex vivo. Four fluorinated DMDPA derivatives were synthesized, two were labeled with fluorine-18: [18F]fluoroethyl-methyldiphenylammonium trifluoromethanesulfonate ([18F]FEMDPA) and [18F]fluorobuthyl-methyldiphenylammonium trifluoromethanesulfonate ([18F]FBMDPA). The other two were labeled using carbon-11: [11C]methyl-(3-fluorophenyl)-methylphenylammonium trifluoromethanesulfonate ([11C]3-F-DMDPA) and [11C]methyl-(4-fluorophenyl)-methylphenylammonium trifluoromethanesulfonate ([11C]4-F-DMDPA). All four DMDPA derivatives exhibited significantly lower heart/liver radioactivity uptake ratios (0.6, 0.4, 0.7 and 0.6, respectively) compared to that of [11C]DMDPA (1.2). Conversely, the two radiolabeled quinolinium salt derivatives, [11C]methylquinolinium iodide ([11C]MeQ) and [18F]FEtQ demonstrated improved heart/liver ratios (2.0 and 1.3, respectively) with clear visualization of the left ventricle myocardium. Renal clearance was the major route of elimination. Among the fluorinated quaternary ammonium salts tested, [18F]FEtQ yielded the best images. Further studies are in progress to elucidate the underlying mechanism of its cardiac uptake.

Rapid Tumor Targeting of Renal-Clearable ZW800-1 Conjugate for Efficient Photothermal Cancer Therapy

The combination of near-infrared (NIR) fluorophores and photothermal therapy (PTT) provides a new opportunity for safe and effective cancer treatment. However, the precise molecular design of functional NIR fluorophores with desired properties, such as high tumor targetability and low nonspecific uptake, remains challenging. In this study, a renal-clearable NIR fluorophore conjugate with high tumor targetability was developed for efficient photothermal cancer therapy. The isoniazid (INH)–ZW800-1 conjugate (INH–ZW) was synthesized by conjugating an antibiotic drug, INH, with a well-known zwitterionic NIR fluorophore, ZW800-1, to improve in vivo performance and fluorescence-guided cancer phototherapy. INH–ZW not only showed rapid tumor accumulation without nonspecific tissue/organ uptake within 1 h after the injection but also generated thermal energy to induce cancer cell death under NIR laser irradiation. Compared with previously reported ZW800-1 conjugates, INH–ZW preserved the ideal biodistribution of ZW800-1 and facilitated improved tumor targeting and PTT. Together, these results demonstrate that the INH–ZW conjugate has great potential to serve as an effective PTT agent capable of rapid tumor targeting and high renal clearance, with excellent photothermal efficacy.

Impact of charge patches on tumor disposition and biodistribution of therapeutic antibodies

This study explores the impact of antibody surface charge on tissue distribution into various tissues including tumor. Tumor-bearing mice were dosed intravenously with a mixture of three antibodies engineered to carry negative charge patches, a balanced charge distribution, or positive patches, respectively. Tissue levels were analyzed with a specific LC-MS/MS method. In addition, the antibody mix was administered to non-tumor bearing mice. Muscle and skin interstitial fluid were obtained by centrifugation and analyzed by LC-MS/MS. An in-vitro endothelium model was explored for its feasibility to mimic the observed distribution differences. A balanced charge distribution was optimal in terms of total tumor exposure, while in other tissues negatively charged and balanced charged antibodies gave similar results. In contrast, positive charge patches generally result in increased serum clearance but markedly enhance tumor and organ uptake, leading to higher tissue-to-serum ratios. The uptake and availability in the interstitial space were confirmed by specific assessment of antibody levels in the interstitial fluid of muscle and skin, with similar charge impact as in total tissue. The in vitro model was able to differentiate the transport propensity of this series of antibody variants. In summary, our results show the differential effects of charge patches on an antibody surface on biodistribution and tumor uptake. These insights may help in the design of molecules with biodistribution properties tailored to their purpose and an optimized safety profile.

Dose predictions for [177Lu]Lu-DOTA-panitumumab F(ab′)2 in NRG mice with HNSCC patient-derived tumour xenografts based on [64Cu]Cu-DOTA-panitumumab F(ab′)2 – implications for a PET theranostic strategy

Background Epidermal growth factor receptors (EGFR) are overexpressed on many head and neck squamous cell carcinoma (HNSCC). Radioimmunotherapy (RIT) with F(ab')2 of the anti-EGFR monoclonal antibody panitumumab labeled with the β-particle emitter, 177Lu may be a promising treatment for HNSCC. Our aim was to assess the feasibility of a theranostic strategy that combines positron emission tomography (PET) with [64Cu]Cu-DOTA-panitumumab F(ab')2 to image HNSCC and predict the radiation equivalent doses to the tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2. Results Panitumumab F(ab')2 were conjugated to DOTA and complexed to 64Cu or 177Lu in high radiochemical purity (95.6 ± 2.1% and 96.7 ± 3.5%, respectively) and exhibited high affinity EGFR binding (Kd = 2.9 ± 0.7 × 10− 9 mol/L). Biodistribution (BOD) studies at 6, 24 or 48 h post-injection (p.i.) of [64Cu]Cu-DOTA-panitumumab F(ab')2 (5.5–14.0 MBq; 50 μg) or [177Lu]Lu-DOTA-panitumumab F(ab')2 (6.5 MBq; 50 μg) in NRG mice with s.c. HNSCC patient-derived xenografts (PDX) overall showed no significant differences in tumour uptake but modest differences in normal organ uptake were noted at certain time points. Tumours were imaged by microPET/CT with [64Cu]Cu-DOTA-panitumumab F(ab')2 or microSPECT/CT with [177Lu]Lu-DOTA-panitumumab F(ab')2 but not with irrelevant [177Lu]Lu-DOTA-trastuzumab F(ab')2. Tumour uptake at 24 h p.i. of [64Cu]Cu-DOTA-panitumumab F(ab')2 [14.9 ± 1.1% injected dose/gram (%ID/g) and [177Lu]Lu-DOTA-panitumumab F(ab')2 (18.0 ± 0.4%ID/g) were significantly higher (P < 0.05) than [177Lu]Lu-DOTA-trastuzumab F(ab')2 (2.6 ± 0.5%ID/g), demonstrating EGFR-mediated tumour uptake. There were no significant differences in the radiation equivalent doses in the tumour and most normal organs estimated for [177Lu]Lu-DOTA-panitumumab F(ab')2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab')2 compared to those estimated directly from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab')2 except for the liver and whole body which were modestly underestimated by [64Cu]Cu-DOTA-panitumumab F(ab')2. Region-of-interest (ROI) analysis of microPET/CT images provided dose estimates for the tumour and liver that were not significantly different for the two radioimmunoconjugates. Human doses from administration of [177Lu]Lu-DOTA-panitumumab F(ab')2 predicted that a 2 cm diameter HNSCC tumour in a patient would receive 1.1–1.5 mSv/MBq and the whole body dose would be 0.15–0.22 mSv/MBq. Conclusion A PET theranostic strategy combining [64Cu]Cu-DOTA-panitumumab F(ab')2 to image HNSCC tumours and predict the equivalent radiation doses in the tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2 is feasible. RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2 may be a promising approach to treatment of HNSCC due to frequent overexpression of EGFR.

Spike protein multiorgan tropism suppressed by antibodies targeting SARS-CoV-2

While there is clinical evidence of severe acute respiratory syndrome coronavirus 2 multiorgan tropism in severely infected coronavirus 19 patients, it is unclear if there is differential multiorgan biodistribution and organ uptake in healthy young individuals, a group that usually has asymptomatic to moderate coronavirus 19 symptoms. In addition, for antibody therapies and vaccines that target the spike protein, it is unclear if these reduce severe acute respiratory syndrome coronavirus 2 or spike protein multiorgan tropism equally. We used fluorescently labeled spike protein near infrared fluorescence to study viral behavior, using an in vivo dynamic imaging system, in young mice. We found a spike protein body-wide biodistribution followed by a slow regional elimination, except for the liver, which showed an accumulation. Spike protein uptake was highest for the lungs, and this was followed by kidney, heart and liver, but, unlike the choroid plexus, it was not detected in the brain parenchyma or cerebrospinal fluid. Thus, the brain vascular barriers were effective in restricting the entry of spike protein into brain parenchyma in young healthy mice. While both anti-angiotensin converting enzyme 2 and anti-spike protein antibodies suppressed spike protein biodistribution and organ uptake, anti-spike protein antibody was more effective. By extension, our data support the efficacy of these antibodies on severe acute respiratory syndrome coronavirus 2 biodistribution kinetics and multiorgan tropism that could determine coronavirus 19 organ-specific outcomes.

Decorating sdAbs with Chelators: Effect of Conjugation on Biodistribution and Functionality

Single domain antibodies (sdAbs) have proven to be valuable probes for molecular imaging. In order to produce such probes, one strategy is the functionalization of the reactive amine side chain of lysines with a chelator, resulting in a mixture of compounds with a different degree of conjugation. In this study, we implemented anion exchange chromatography (AEX) to separate the different compounds or fractions that were further characterized and evaluated to study the impact of the conjugation degree on pharmacokinetic properties and functionality. Anti-HER2 and anti-MMR sdAbs were functionalized with NOTA or DTPA chelator. Anion exchange chromatography was performed using 0.02 mol/L Tris pH 7.5 as the first solvent and 0.25 M or 0.4 M NaCl (in case of NOTA chelator or DTPA chelator, respectively) as the second solvent applied as a gradient. The fractions were characterized via mass spectrometry (MS), surface plasmon resonance (SPR), and isoelectric focusing gel electrophoresis (IEF), while in vivo studies were performed after radiolabeling with either 68Ga (NOTA) or 111In (DTPA) to assess the impact of the conjugation degree on pharmacokinetics. AEX could successfully be applied to separate fractions of (chelator)n-anti-HER2 and (chelator)n-anti-MMR sdAb constructs. MS confirmed the identity of different peaks obtained in the separation process. SPR measurement suggests a small loss of affinity for (chelator)3-anti-sdAb, while IEF revealed a correlated decrease in isoelectric point (pI) with the number of conjugated chelators. Interestingly, both the reduction in affinity and in pI was stronger with the DTPA chelator than with NOTA for both sdAbs. In vivo data showed no significant differences in organ uptake for any construct, except for (DTPA)n-anti-MMR, which showed a significantly higher liver uptake for (DTPA)1-anti-MMR compared to (DTPA)2-anti-MMR and (DTPA)3-anti-MMR. For all constructs in general, high kidney uptake was observed, due to the typical renal clearance of sdAb-based tracers. The kidney uptake showed significant differences between fractions of a same construct and indicates that a higher conjugation degree improves kidney clearance. AEX allows the separation of sdAbs with a different degree of conjugation and provides the opportunity to further characterize individual fractions. The conjugation of a chelator to sdAbs can alter some properties of the tracers, such as pI; however, the impact on the general biodistribution profile and tumor targeting was minimal.

Equivalent tumor detection for early and late FAPI-46 PET acquisition

Introduction Positron emission tomography (PET) using small ligands of the fibroblast activation protein (FAP) was recently introduced. However, optimal uptake time has not been defined yet. Here, we systematically compare early (~ 10 min p.i.) and late (~ 60 min p.i.) FAPI-46 imaging in patients with various types of cancer. Methods This is a retrospective single-institutional study. Imaging was performed at the Essen University Hospital, Germany. A total of 69 patients who underwent dual time-point imaging for either restaging (n = 52, 75%) or staging (n = 17, 25%) of cancer were included. Patients underwent PET with two acquisitions: early (mean 11 min, SD 4) and late (mean 66 min, SD 9). Mean injected activity was 148 MBq (SD 33). Results In total, 400 lesions were detected in 69 patients. Two of 400 (0.5%) lesions were only seen in early time-point imaging but not in late time-point imaging. On a per-patient level, there was no significant difference between SUVmax of hottest tumor lesions (Wilcoxon: P = 0.73). Organ uptake demonstrated significant early to late decrease in SUVmean (average ∆SUVmean: − 0.48, − 0.14, − 0.27 for gluteus, liver, and mediastinum, respectively; Wilcoxon: P < 0.001). On a per-lesion basis, a slight increase of SUVmax was observed (average ∆SUVmax: + 0.4, Wilcoxon: P = 0.03). Conclusion In conclusion, early (~ 10 min p.i.) versus late (~ 60 min p.i.) FAPI-46 imaging resulted in equivalent lesion uptake and tumor detection. For improved feasibility and scan volume, we implement early FAPI-46 PET in future clinical and research protocols.

66Ga-PET-imaging of GRPR-expression in prostate cancer: production and characterization of [66Ga]Ga-NOTA-PEG2-RM26

Molecular imaging of the gastrin-releasing peptide receptor (GRPR) could improve patient management in prostate cancer. This study aimed to produce gallium-66 (T½ = 9.5 h) suitable for radiolabeling, and investigate the imaging properties of gallium-66 labeled GRPR-antagonist NOTA-PEG2-RM26 for later-time point PET-imaging of GRPR expression. Gallium-66 was cyclotron-produced using a liquid target, and enriched [66Zn]Zn(NO3)2. In vitro, [66Ga]Ga-NOTA-PEG2-RM26 was characterized in GRPR-expressing PC-3 prostate cancer cells. In vivo, specificity test and biodistribution studies were performed 3 h and 22 h pi in PC-3 xenografted mice. microPET/MR was performed 3 h and 22 h pi. Biodistribution of [66Ga]Ga-NOTA-PEG2-RM26 was compared with [68Ga]Ga-NOTA-PEG2-RM26 3 h pi. [66Ga]Ga-NOTA-PEG2-RM26 was successfully prepared with preserved binding specificity and high affinity towards GRPR. [66Ga]Ga-NOTA-PEG2-RM26 cleared rapidly from blood via kidneys. Tumor uptake was GRPR-specific and exceeded normal organ uptake. Normal tissue clearance was limited, resulting in no improvement of tumor-to-organ ratios with time. Tumors could be clearly visualized using microPET/MR. Gallium-66 was successfully produced and [66Ga]Ga-NOTA-PEG2-RM26 was able to clearly visualize GRPR-expression both shortly after injection and on the next day using PET. However, delayed imaging did not improve contrast for Ga-labeled NOTA-PEG2-RM26.