Jonas Rosager Henriksen

Background: Antibiotic-loaded ceramic bone grafts (ACBGs) are widely assumed to provide extended local antimicrobial protection, maintaining concentrations above the minimum inhibitory concentration (MIC) for 28 to 42 days. However, this time frame is mainly based on in vitro elution assays with simplified washout models, and its relevance to in vivo performance is uncertain. This study compared the in vivo and in vitro release and behavior of gentamicin from 2 commercial ACBGs. Methods: Gentamicin-loaded hydroxyapatite/calcium sulfate (HA-CS) and calcium sulfate (CS) composites were investigated using a murine implantation model and in vitro elution assays. Gentamicin content in recovered depot material and elution media was quantified by liquid chromatography with tandem mass spectrometry. Model calculations evaluated how the assay design affects the estimated time above the MIC. Results: Both ACBGs released >98% of gentamicin within 6 hours and >99% within 24 hours in vivo. In vitro assays likewise indicated early depot depletion, with concentration-time profiles beyond 48 hours driven by serial dilution rather than release. Model calculations suggested that the apparent prolonged antibiotic activity in previous in vitro assays was largely due to the assay design, not to sustained release, implying that freely administered antibiotics could produce similar profiles. Conclusions: Neither ACBG demonstrated measurable sustained release beyond an initial 24-hour burst phase. This may have important clinical implications, as expectations of prolonged local antimicrobial protection are not supported by the cumulative-release data showing rapid antibiotic depot depletion.

Brachytherapy (BT) is routinely used in the treatment of various cancers. Current BT relies on the placement of large sources of radioactivity at the tumor site, requiring applicators that may cause local traumas and lesions. Further, they suffer from inflexibility in where they can be placed and some sources reside permanently in the body, causing potential long-term discomfort. These issues can be circumvented through injectable sources, prepared as biodegradable materials containing radionuclides that form solid seeds after administration. The level of radioactivity contained in such seeds must be sufficient to achieve substantial local irradiation. In this report, we investigate two different strategies for biodegradable BT seeds. The first strategy entails injectable seeds based on 103Pd-labeled palladium-gold alloy nanoparticles ([103Pd]PdAuNPs). These were prepared by combining [103Pd]PdH2Cl4 and AuHCl4, followed by lipophilic surface coating and dispersed in lactose octaisobutyrate and ethanol (LOIB:EtOH), in overall radiochemical yield (RCY) of 83%. With the second strategy, [103Pd]Pd-SSIB was prepared by conjugating the [16]aneS4 chelator with lipophilic sucrose septaisobutyrate (SSIB) followed by complexation with [103Pd]PdH2Cl4 (RCY = 99%) and mixed with LOIB:EtOH. [103Pd]Pd-SSIB was likewise formulated as injectable liquid forming seeds by mixing with LOIB. Both formulations reached activities of 1.0–1.5 GBq/mL and negligible release of radioactivity after injection of 100 µL (100–150 MBq) into aqueous buffer or mouse serum of less than 1% over one month. Both strategies for forming injectable BT seeds containing high 103Pd activity resulted in high radiolabeling yields, high activity per seed, and high activity retention. We consider both strategies suitable for BT, with the preferable strategy using a [16]aneS4 chelator due to its higher biodegradability.

INKT cells – often referred to as the “Swiss Army knife” of the immune system – have emerged as central players in cancer vaccine therapies.

Background: The accumulation of liposome encapsulated chemotherapy in solid cancers is dependent on the presence of the enhanced permeability and retention (EPR) effect. Positron emission tomography (PET) imaging with a liposome encapsulated radioisotope, such as liposome encapsulated Cu-64 (⁶⁴Cu-liposome) may help to identify tumors with high liposome accumulation, and thereby stratify patients based on expected benefit from liposomal chemotherapy. However, intravenous administration of liposomes without a cytotoxic content is complicated by the accelerated blood clearance (ABC) phenomenon for succeeding therapeutic liposome dosing. Alternative markers for assessing the tumor’s EPR level are therefore warranted. Materials and Methods: To increase our understanding of EPR variations and to ultimately identify an alternative marker for the EPR effect, we investigated the correlation between ⁶⁴Cu-liposome PET/CT (EPR effect) and ⁶⁸Ga-RGD PET/CT (neoangiogenesis), ¹⁸F-FDG PET/CT (glycolysis), diffusion-weighted MRI (diffusivity) and interstitial fluid pressure in two experimental cancer models (CT26 and COLO 205). Results: ⁶⁴Cu-liposome and ⁶⁸Ga-RGD SUV_max displayed a significant moderate correlation, however, none of the other parameters evaluated displayed significant correlations. These results indicate that differences in neoangiogenesis may explain some EPR variability, however, as correlations were only moderate and not observed for SUV_mean, ⁶⁸Ga-RGD is probably insufficient to serve as a stand-alone surrogate marker for quantifying the EPR effect and stratifying patients.

The cations, Cu ²⁺ , Zn ²⁺ , Ag ⁺ and Au ³⁺ display anomalous membrane permeability, which is utilized for synthesis of gold nanoparticles inside liposomes.

The liquid-based Carbo-gel technology enables real-time multimodal image guidance for improved therapeutic precision.

Background: Active, ligand-mediated, targeting of functionalized liposomes to folate receptors (FRs) overexpressed on cancer cells could potentially improve drug delivery and specificity. Studies on folate-targeting liposomes (FTLs) have, however, yielded varying results and generally fail to display a clear benefit of FR targeting. Method: Tumor accumulating potential of FTLs and NTLs were investigated in a FR overexpressing xenograft model by positron emission tomography/computed tomography imaging. Results: Tumors displayed significantly lower activity of FTLs than NTLs. Furthermore, FTLs displayed worse circulating properties and increased liver-accumulation than NTLs. Conclusion: This study underlines that long-circulating properties of liposomes must be achieved to take advantage of EPR-dependent tumor accumulation which may be lost by functionalization. FR-functionalization negatively affected both tumor accumulation and circulation properties.

The enhanced permeability and retention (EPR) effect increases tumor accumulation of liposomal chemotherapy and should, in theory, increase anticancer effects and lower toxicity. Unfortunately, liposomal chemotherapy has generally not met the expected potential, perhaps because the EPR effect is not ubiquitous. PET imaging using radiolabeled liposomes can identify cancers positive for the EPR effect. In the current study, we show in clinical canine cancer patients that repeated imaging with radiolabeled liposomes (64Cu-liposome) induces the accelerated blood clearance (ABC) phenomenon. This was observed even with very long intervals between liposome injections, which contradict previous reporting in experimental animal models. The induction of ABC may be devastating for the theranostic use of liposomal imaging, as this could vaccinate patients against therapeutic efficacy. To investigate and solve this important problem, an additional study part was designed in which rats were subjected to repeated liposomal administrations, including stealth 64Cu-liposome PET imaging and Caelyx® chemotherapy. Most importantly, it was found that by increasing the lipid dose at the first injection or by supplying a small pre-dose before the second 64Cu-liposome injection, ABC could be prevented. Importantly, signs of liposome tracer breakdown with subsequent renal excretion were observed. These findings highlight the importance of the ABC phenomenon for liposomal predictive imaging in a clinically relevant setting and show that carefully planned application is central to avoid potential detrimental effects on patient benefit.

Long circulating liposomes entrapping iodinated and radioiodinated compounds offer a highly versatile theranostic platform. Here we report a new methodology for efficient and high-yield loading of such compounds into liposomes, enabling CT/SPECT/PET imaging and ¹³¹I-radiotherapy.