Highlights d We have developed a trained immunity-inducing nanobiologic therapeutic named MTP-HDL d MTP-HDL favorably accumulates in hematopoietic organs of mice and non-human primates d MTP-HDL nanotherapy induces trained immunity through bone marrow progenitors in vivo d MTP-HDL nanotherapy inhibits tumor growth and potentiates immune checkpoint inhibition

Nanomedicine research produces hundreds of studies every year, yet very few formulations have been approved for clinical use. This is due in part to a reliance on murine studies, which have limited value in accurately predicting translational efficacy in larger animal models and humans. Here, we report the scale-up of a nanoimmunotherapy from mouse to large rabbit and porcine atherosclerosis models, with an emphasis on the solutions we implemented to overcome production and evaluation challenges. Specifically, we integrated translational imaging readouts within our workflow to both analyze the nanoimmunotherapeutic’s in vivo behavior and assess treatment response in larger animals. We observed our nanoimmunotherapeutic’s anti-inflammatory efficacy in mice, as well as rabbits and pigs. Nanoimmunotherapy-mediated reduction of inflammation in the large animal models halted plaque progression, supporting the approach’s translatability and potential to acutely treat atherosclerosis.

Although the first nanomedicine was clinically approved more than two decades ago, nanoparticles’ (NP) in vivo behavior is complex and the immune system’s role in their application remains elusive. At present, only passive-targeting nanoformulations have been clinically approved, while more complicated active-targeting strategies typically fail to advance from the early clinical phase stage. This absence of clinical translation is, among others, due to the very limited understanding for in vivo targeting mechanisms. Dynamic in vivo phenomena such as NPs’ real-time targeting kinetics and phagocytes’ contribution to active NP targeting remain largely unexplored. To better understand in vivo targeting, monitoring NP accumulation and distribution at complementary levels of spatial and temporal resolution is imperative. Here, we integrate in vivo positron emission tomography/computed tomography imaging with intravital microscopy and flow cytometric analyses to study α v β 3 -integrin-targeted cyclic arginine-glycine-aspartate decorated liposomes and oil-in-water nanoemulsions in tumor mouse models. We observed that ligand-mediated accumulation in cancerous lesions is multifaceted and identified “NP hitchhiking” with phagocytes to contribute considerably to this intricate process. We anticipate that this understanding can facilitate rational improvement of nanomedicine applications and that immune cell–NP interactions can be harnessed to develop clinically viable nanomedicine-based immunotherapies.

This study aimed to (i) develop Magnetization-Prepared Golden-angle RAdial Sparse Parallel (MP-GRASP) MRI using a stack-of-stars trajectory for rapid free-breathing T1 mapping and (ii) extend MP-GRASP to multi-echo acquisition (MP-Dixon-GRASP) for fat/water-separated (water-specific) T1 mapping. Methods: An adiabatic non-selective 180° inversion-recovery pulse was added to a gradient-echo-based golden-angle stack-of-stars sequence for magnetizationprepared 3D single-echo or 3D multi-echo acquisition. In combination with subspace-based GRASP-Pro reconstruction, the sequence allows for standard T1 mapping (MP-GRASP) or fat/water-separated T1 mapping (MP-Dixon-GRASP), respectively. The accuracy of T1 mapping using MP-GRASP was evaluated in a phantom and volunteers (brain and liver) against clinically accepted reference methods. The repeatability of T1 estimation was also assessed in the phantom and volunteers. The performance of MP-Dixon-GRASP for water-specific T1 mapping was evaluated in a fat/water phantom and volunteers (brain and liver). Results: ROI-based mean T1 values are correlated between the references and MP-GRASP in the phantom (R 2 = 1.0), brain (R 2 = 0.96), and liver (R 2 = 0.73). MP-GRASP achieved good repeatability of T1 estimation in the phantom (R 2 = 1.0), brain (R 2 = 0.99), and liver (R 2 = 0.82). Water-specific T1 is different from in-phase and out-of-phase composite T1 (composite T1 when fat and water signal are mixed in phase or out of phase) both in the phantom and volunteers. Conclusion: This work demonstrated the initial performance of MP-GRASP and MP-Dixon-GRASP MRI for rapid 3D T1 mapping and 3D fat/water-separated T1 mapping in the brain (without motion) and in the liver (during free breathing). With fat/water-separated T1 estimation, MP-Dixon-GRASP could be potentially useful for imaging patients with fatty-liver diseases.

Acute bacterial endocarditis is a rapid, difficult to manage, and frequently lethal disease. Potent antibiotics often cannot efficiently kill Staphylococcus aureus that colonizes the heart’s valves. S. aureus relies on virulence factors to evade therapeutics and the host’s immune response, usurping the host’s clotting system by activating circulating prothrombin with staphylocoagulase and von Willebrand factor–binding protein. An insoluble fibrin barrier then forms around the bacterial colony, shielding the pathogen from immune cell clearance. Targeting virulence factors may provide previously unidentified avenues to better diagnose and treat endocarditis. To tap into this unused therapeutic opportunity, we codeveloped therapeutics and multimodal molecular imaging to probe the host-pathogen interface. We introduced and validated a family of small-molecule optical and positron emission tomography (PET) reporters targeting active thrombin in the fibrin-rich environment of bacterial colonies. The imaging agents, based on the clinical thrombin inhibitor dabigatran, are bound to heart valve vegetations in mice. Using optical imaging, we monitored therapy with antibodies neutralizing staphylocoagulase and von Willebrand factor–binding protein in mice with S. aureus endocarditis. This treatment deactivated bacterial defenses against innate immune cells, decreased in vivo imaging signal, and improved survival. Aortic or tricuspid S. aureus endocarditis in piglets was also successfully imaged with clinical PET/magnetic resonance imaging. Our data map a route toward adjuvant immunotherapy for endocarditis and provide efficient tools to monitor this drug class for infectious diseases.

Immunotherapies controlling the adaptive immune system are firmly established, but regulating the innate immune system remains much less explored. The intrinsic interactions between nanoparticles and phagocytic myeloid cells make these materials especially suited for engaging the innate immune system. However, developing nanotherapeutics is an elaborate process. Here, we demonstrate a modular approach that facilitates efficiently incorporating a broad variety of drugs in a nanobiologic platform. Using a microfluidic formulation strategy, we produced apolipoprotein A1–based nanobiologics with favorable innate immune system–engaging properties as evaluated by in vivo screening. Subsequently, rapamycin and three small-molecule inhibitors were derivatized with lipophilic promoieties, ensuring their seamless incorporation and efficient retention in nanobiologics. A short regimen of intravenously administered rapamycin-loaded nanobiologics (mTORi-NBs) significantly prolonged allograft survival in a heart transplantation mouse model. Last, we studied mTORi-NB biodistribution in nonhuman primates by PET/MR imaging and evaluated its safety, paving the way for clinical translation.

Significant improvements in radiotherapy are likely to come from biological rather than technical optimization, for example increasing tumour radiosensitivity via combination with targeted therapies. Such paradigms must first be evaluated in preclinical models for efficacy, and recent advances in small animal radiotherapy research platforms allow advanced irradiation protocols, similar to those used clinically, to be carried out in orthotopic models. Dose assessment in such systems is complex however, and a lack of established tools and methodologies for traceable and accurate dosimetry is currently limiting the capabilities of such platforms and slowing the clinical uptake of new approaches. Here we report the creation of an anatomically correct phantom, fabricated from materials with tissue-equivalent electron density, into which dosimetry detectors can be incorporated for measurement as part of quality control (QC). The phantom also allows training in preclinical radiotherapy planning and cross-institution validation of dose delivery protocols for small animal radiotherapy platforms without the need to sacrifice animals, with high reproducibility. Mouse CT data was acquired and segmented into soft tissue, bone and lung. The skeleton was fabricated using 3D printing, whilst lung was created using computer numerical control (CNC) milling. Skeleton and lung were then set into a surfacerendered mould and soft tissue material added to create a whole-body phantom. Materials for fabrication were characterized

Despite well-established dosimetry in clinical radiotherapy, dose measurements in pre-clinical and radiobiology studies are frequently inadequate, thus undermining the reliability and reproducibility of published findings. The lack of suitable dosimetry protocols, coupled with the increasing complexity of pre-clinical irradiation platforms, undermines confidence in preclinical studies and represents a serious obstacle in the translation to clinical practice. To accurately measure output of a pre-clinical radiotherapy unit, appropriate Codes of Practice (CoP) for medium energy x-rays needs to be employed. However, determination of absorbed dose to water (Dw) relies on application of backscatter factor (Bw) employing in-air method or carrying out in-phantom measurement at the reference depth of 2 cm in a full backscatter (i.e. 30 × 30 × 30 cm3) condition. Both of these methods require thickness of at least 30 cm of underlying material, which are never fulfilled in typical pre-clinical irradiations. This work is focused on evaluation the effects of the lack of recommended reference conditions in dosimetry measurements for pre-clinical settings and is aimed at extending the recommendations of the current CoP to practical experimental conditions and highlighting the potential impact of the lack of correct backscatter considerations on radiobiological studies.