High-Z Metal-Organic Frameworks for X-ray Radiation-Based Cancer Theranostics.

X-ray radiation is commonly employed in clinical practice for diagnostic and therapeutic applications. Over the past decade, developments in nanotechnology have led to the use of high-Z elements as the basis for innovative new treatment platforms that enhance the clinical efficacy of X-ray radiation. Nanoscale metal-frameworks (nMOFs) are coordination networks containing organic ligands that have attracted attention as therapeutic platforms in oncology and other areas of medicine. In cancer therapy, X-ray activated, high-Z nMOFs have demonstrated potential as radiosensitizers that increase local radiation dose deposition and generation of reactive oxygen species (ROS). This minireview summarizes current research on high-Z nMOFs in cancer theranostics and discusses factors that may influence future clinical application.

Lipid Nanoparticle-Delivered Chemically Modified mRNA Restores Chloride Secretion in Cystic Fibrosis.

The promise of gene therapy for the treatment of cystic fibrosis has yet to be fully clinically realized despite years of effort toward correcting the underlying genetic defect in the cystic fibrosis transmembrane conductance regulator (CFTR). mRNA therapy via nanoparticle delivery represents a powerful technology for the transfer of genetic material to cells with large, widespread populations, such as airway epithelia. We deployed a clinically relevant lipid-based nanoparticle (LNP) for packaging and delivery of large chemically modified CFTR mRNA (cmCFTR) to patient-derived bronchial epithelial cells, resulting in an increase in membrane-localized CFTR and rescue of its primary function as a chloride channel. Furthermore, nasal application of LNP-cmCFTR restored CFTR-mediated chloride secretion to conductive airway epithelia in CFTR knockout mice for at least 14 days. On day 3 post-transfection, CFTR activity peaked, recovering up to 55% of the net chloride efflux characteristic of healthy mice. This magnitude of response is superior to liposomal CFTR DNA delivery and is comparable with outcomes observed in the currently approved drug ivacaftor. LNP-cmRNA-based systems represent a powerful platform technology for correction of cystic fibrosis and other monogenic disorders.

Rare-Earth-Doped Nanoparticles for Short-Wave Infrared Fluorescence Bioimaging and Molecular Targeting of αVß3-Expressing Tumors.

The use of short-wave infrared (SWIR) light for fluorescence bioimaging offers the advantage of reduced photon scattering and improved tissue penetration compared to traditional shorter wavelength imaging approaches. While several nanomaterials have been shown capable of generating SWIR emissions, rare-earth-doped nanoparticles (REs) have emerged as an exceptionally bright and biocompatible class of SWIR emitters. Here, we demonstrate SWIR imaging of REs for several applications, including lymphatic mapping, real-time monitoring of probe biodistribution, and molecular targeting of the αvß3 integrin in a tumor model. We further quantified the resolution and depth penetration limits of SWIR light emitted by REs in a customized imaging unit engineered for SWIR imaging of live small animals. Our results indicate that SWIR light has broad utility for preclinical biomedical imaging and demonstrates the potential for molecular imaging using targeted REs.

Boosting Intracellular Delivery of Lipid Nanoparticle-Encapsulated mRNA.

Intracellular delivery of mRNA holds great potential for vaccine1-3 and therapeutic4 discovery and development. Despite increasing recognition of the utility of lipid-based nanoparticles (LNPs) for intracellular delivery of mRNA, particle engineering is hindered by insufficient understanding of endosomal escape, which is believed to be a main limiter of cytosolic availability and activity of the nucleic acid inside the cell. Using a series of CRISPR-based genetic perturbations of the lysosomal pathway, we have identified that late endosome/lysosome (LE/Ly) formation is essential for functional delivery of exogenously presented mRNA. Lysosomes provide a spatiotemporal hub to orchestrate mTOR signaling and are known to control cell proliferation, nutrient sensing, ribosomal biogenesis, and mRNA translation. Through modulation of the mTOR pathway we were able to enhance or inhibit LNP-mediated mRNA delivery. To further boost intracellular delivery of mRNA, we screened 212 bioactive lipid-like molecules that are either enriched in vesicular compartments or modulate cell signaling. Surprisingly, we have discovered that leukotriene-antagonists, clinically approved for treatment of asthma and other lung diseases, enhance intracellular mRNA delivery in vitro (over 3-fold, p < 0.005) and in vivo (over 2-fold, p < 0.005). Understanding LNP-mediated intracellular delivery will inspire the next generation of RNA therapeutics that have high potency and limited toxicity.

X-ray-induced shortwave infrared biomedical imaging using rare-earth nanoprobes.

Shortwave infrared (SWIR or NIR-II) light provides significant advantages for imaging biological structures due to reduced autofluorescence and photon scattering. Here, we report on the development of rare-earth nanoprobes that exhibit SWIR luminescence following X-ray irradiation. We demonstrate the ability of X-ray-induced SWIR luminescence (X-IR) to monitor biodistribution and map lymphatic drainage. Our results indicate X-IR imaging is a promising new modality for preclinical applications and has potential for dual-modality molecular disease imaging.

Efficient Radioisotope Energy Transfer by Gold Nanoclusters for Molecular Imaging.

Beta-emitting isotopes Fluorine-18 and Yttrium-90 are tested for their potential to stimulate gold nanoclusters conjugated with blood serum proteins (AuNCs). AuNCs excited by either medical radioisotope are found to be highly effective ionizing radiation energy transfer mediators, suitable for in vivo optical imaging. AuNCs synthesized with protein templates convert beta-decaying radioisotope energy into tissue-penetrating optical signals between 620 and 800 nm. Optical signals are not detected from AuNCs incubated with Technetium-99m, a pure gamma emitter that is used as a control. Optical emission from AuNCs is not proportional to Cerenkov radiation, indicating that the energy transfer between the radionuclide and AuNC is only partially mediated by Cerenkov photons. A direct Coulombic interaction is proposed as a novel and significant mechanism of energy transfer between decaying radionuclides and AuNCs.

Synergistic assembly of heavy metal clusters and luminescent organic bridging ligands in metal-organic frameworks for highly efficient X-ray scintillation.

We have designed two metal-organic frameworks (MOFs) to efficiently convert X-ray to visible-light luminescence. The MOFs are constructed from M6(µ3-O)4(µ3-OH)4(carboxylate)12 (M = Hf or Zr) secondary building units (SBUs) and anthracene-based dicarboxylate bridging ligands. The high atomic number of Zr and Hf in the SBUs serves as effective X-ray antenna by absorbing X-ray photons and converting them to fast electrons through the photoelectric effect. The generated electrons then excite multiple anthracene-based emitters in the MOF through inelastic scattering, leading to efficient generation of detectable photons in the visible spectrum. The MOF materials thus serve as efficient X-ray scintillators via synergistic X-ray absorption by the metal-cluster SBUs and optical emission by the bridging ligands.

Nanoscale Hafnium Metal-Organic Frameworks Enhance Radiotherapeutic Effects by Upregulation of Type I Interferon and TLR7 Expression.

Recent preclinical and clinical studies have highlighted the improved outcomes of combination radiotherapy and immunotherapy. Concurrently, the development of high-Z metallic nanoparticles as radiation dose enhancers has been explored to widen the therapeutic window of radiotherapy and potentially enhance immune activation. In this study, folate-modified hafnium-based metal-organic frameworks (HfMOF-PEG-FA) are evaluated in combination with imiquimod, a TLR7 agonist, as a well-defined interferon regulatory factor (IRF) stimulator for local antitumor immunotherapy. The enhancement of radiation dose deposition by HfMOF-PEG-FA and subsequent generation of reactive oxygen species (ROS) deregulates cell proliferation and increases apoptosis. HfMOF-PEG-FA loaded with imiquimod (HfMOF-PEG-FA@IMQ) increases DNA double-strand breaks and cell death, including apoptosis, necrosis, and calreticulin exposure, in response to X-ray irradiation. Treatment with this multipronged therapy promotes IRF stimulation for subsequent interferon production within tumor cells themselves. The novel observation is reported that HfMOF itself increases TLR7 expression, unexpectedly pairing immune agonist and receptor upregulation in a tumor intrinsic manner, and supporting the synergistic effect observed with the γH2AX assay. T-cell analysis of CT26 tumors following intratumoral administration of HfMOF-PEG-FA@IMQ with radiotherapy reveals a promising antitumor response, characterized by an increase in CD8+ and proliferative T cells.

Peptide-guided lipid nanoparticles deliver mRNA to the neural retina of rodents and nonhuman primates.

Lipid nanoparticle (LNP)-based mRNA delivery holds promise for the treatment of inherited retinal degenerations. Currently, LNP-mediated mRNA delivery is restricted to the retinal pigment epithelium (RPE) and Müller glia. LNPs must overcome ocular barriers to transfect neuronal cells critical for visual phototransduction, the photoreceptors (PRs). We used a combinatorial M13 bacteriophage-based heptameric peptide phage display library for the mining of peptide ligands that target PRs. We identified the most promising peptide candidates resulting from in vivo biopanning. Dye-conjugated peptides showed rapid localization to the PRs. LNPs decorated with the top-performing peptide ligands delivered mRNA to the PRs, RPE, and Müller glia in mice. This distribution translated to the nonhuman primate eye, wherein robust protein expression was observed in the PRs, Müller glia, and RPE. Overall, we have developed peptide-conjugated LNPs that can enable mRNA delivery to the neural retina, expanding the utility of LNP-mRNA therapies for inherited blindness.

Radiotherapy reimagined: Integrating nanomedicines into radiotherapy clinical trials.

Radioenhancing nanoparticles (NPs) are being evaluated in ongoing clinical trials for various cancers including head and neck, lung, esophagus, pancreas, prostate, and soft tissue sarcoma. Supported by decades of preclinical investigation and recent randomized trial data establishing clinical activity, these agents are poised to influence future multimodality treatment paradigms involving radiotherapy. Although the physical interactions between NPs and ionizing radiation are well characterized, less is known about how these agents modify the tumor microenvironment, particularly regarding tumor immunogenicity. In this review, we describe the key multidisciplinary considerations related to radiation, surgery, immunology, and pathology for designing radioenhancing NP clinical trials. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease.

Radioluminescent nanophosphors enable multiplexed small-animal imaging.

We demonstrate the ability to image multiple nanoparticle-based contrast agents simultaneously using a nanophosphor platform excited by either radiopharmaceutical or X-ray irradiation. These radioluminescent nanoparticles emit optical light at unique wavelengths depending on their lanthanide dopant, enabling multiplexed imaging. This study demonstrates the separation of two distinct nanophosphor contrast agents in gelatin phantoms with a recovered phosphor separation correlation of -0.98. The ability to distinguish the two nanophosphors and a Cerenkov component is then demonstrated in a small animal phantom. Combined with the high-resolution potential of low-scattering X-ray excitation, this imaging technique may be a promising method to probe molecular processes in living organisms.

Development of a G2/M arrest high-throughput screening method identifies potent radiosensitizers.

Radiation is a powerful tool used to control tumor growth and induce an immune response; however, it is limited by damage to surrounding tissue and adverse effects such skin irritation. Breast cancer patients in particular may endure radiation dermatitis, and potentially lymphedema, after a course of radiotherapy. Radio-sensitizing small molecule drugs may enable lower effective doses of both radiation and chemotherapy to minimize toxicity to healthy tissue. In this study, we identified a novel high-throughput method for screening radiosensitizers by image analysis of nuclear size and cell cycle. In vitro assays were performed on cancer cells lines to assess combined therapeutic and radiation effects. In vivo, radiation in combination with proflavine hemisulfate led to enhanced efficacy demonstrated by improved tumor volume control in mice bearing syngeneic breast tumors. This study provides a proof of concept for utilizing G2/M stall as a predictor of radiosensitization and is the first report of a flavin acting as an X-ray radiation enhancer in a breast cancer mouse model.

An Advanced Thermal Decomposition Method to Produce Magnetic Nanoparticles with Ultrahigh Heating Efficiency for Systemic Magnetic Hyperthermia.

Due to the limited heating efficiency of available magnetic nanoparticles, it is difficult to achieve therapeutic temperatures above 44 °C in relatively inaccessible tumors during magnetic hyperthermia following systemic administration of nanoparticles at clinical dosage (≤10 mg kg-1 ). To address this, a method for the preparation of magnetic nanoparticles with ultrahigh heating capacity in the presence of an alternating magnetic field (AMF) is presented. The low nitrogen flow rate of 10 mL min-1 during the thermal decomposition reaction results in cobalt-doped nanoparticles with a magnetite (Fe3 O4 ) core and a maghemite (γ-Fe2 O3 ) shell that exhibit the highest intrinsic loss power reported to date of 47.5 nH m2 kg-1 . The heating efficiency of these nanoparticles correlates positively with increasing shell thickness, which can be controlled by the flow rate of nitrogen. Intravenous injection of nanoparticles at a low dose of 4 mg kg-1 elevates intratumoral temperatures to 50 °C in mice-bearing subcutaneous and metastatic cancer grafts during exposure to AMF. This approach can also be applied to the synthesis of other metal-doped nanoparticles with core-shell structures. Consequently, this method can potentially be used for the development of novel nanoparticles with high heating performance, further advancing systemic magnetic hyperthermia for cancer treatment.

Glypican-3 targeting of liver cancer cells using multifunctional nanoparticles.

Imaging is essential in accurately detecting, staging, and treating primary liver cancer (hepatocellular carcinoma [HCC]), one of the most prevalent and lethal malignancies. We developed a novel multifunctional nanoparticle (NP) specifically targeting glypican-3 (GPC3), a proteoglycan implicated in promotion of cell growth that is overexpressed in most HCCs. Quantitative real-time polymerase chain reaction was performed to confirm the differential GPC3 expression in two human HCC cells, Hep G2 (high) and HLF (negligible). These cells were treated with biotin-conjugated GPC3 monoclonal antibody (αGPC3) and subsequently targeted using superparamagnetic iron oxide NPs conjugated to streptavidin and Alexa Fluor 647. Flow cytometry demonstrated that only GPC3-expressing Hep G2 cells were specifically targeted using this αGPC3-NP conjugate (fourfold mean fluorescence over nontargeted NP), and magnetic resonance imaging (MRI) experiments showed similar findings (threefold R2 relaxivity). Confocal fluorescence microscopy localized the αGPC3 NPs only to the cell surface of GPC3-expressing Hep G2 cells. Further characterization of this construct demonstrated a negatively charged, monodisperse, 50 nm NP, ideally suited for tumor targeting. This GPC3-specific NP system, with dual-modality imaging capability, may enhance pretreatment MRI, enable refined intraoperative HCC visualization by near-infrared fluorescence, and be potentially used as a carrier for delivery of tumor-targeted therapies, improving patient outcomes.

RNA-Based Therapeutics: Current Developments in Targeted Molecular Therapy of Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a highly heterogeneous and aggressive cancer that has the highest mortality rate out of all breast cancer subtypes. Conventional clinical treatments targeting ER, PR, and HER2 receptors have been unsuccessful in the treatment of TNBC, which has led to various research efforts in developing new strategies to treat TNBC. Targeted molecular therapy of TNBC utilizes knowledge of key molecular signatures of TNBC that can be effectively modulated to produce a positive therapeutic response. Correspondingly, RNA-based therapeutics represent a novel tool in oncology with their ability to alter intrinsic cancer pathways that contribute to poor patient prognosis. Current RNA-based therapeutics exist as two major areas of investigation-RNA interference (RNAi) and RNA nanotherapy, where RNAi utilizes principles of gene silencing, and RNA nanotherapy utilizes RNA-derived nanoparticles to deliver chemotherapeutics to target cells. RNAi can be further classified as therapeutics utilizing either small interfering RNA (siRNA) or microRNA (miRNA). As the broader field of gene therapy has advanced significantly in recent years, so too have efforts in the development of effective RNA-based therapeutic strategies for treating aggressive cancers, including TNBC. This review will summarize key advances in targeted molecular therapy of TNBC, describing current trends in treatment using RNAi, combination therapies, and recent efforts in RNA immunotherapy, utilizing messenger RNA (mRNA) in the development of cancer vaccines.

Exploiting Phagocytic Checkpoints in Nanomedicine: Applications in Imaging and Combination Therapies.

Recent interest in cancer immunotherapy has largely been focused on the adaptive immune system, particularly adoptive T-cell therapy and immune checkpoint blockade (ICB). Despite improvements in overall survival and progression-free survival across multiple cancer types, neither cell-based therapies nor ICB results in durable disease control in the majority of patients. A critical component of antitumor immunity is the mononuclear phagocyte system and its role in both innate and adaptive immunity. The phagocytic functions of these cells have been shown to be modulated through multiple pathways, including the CD47-SIRPα axis, which is manipulated by cancer cells for immune evasion. In addition to CD47, tumors express a variety of other "don't eat me" signals, including beta-2-microglobulin and CD24, and "eat me" signals, including calreticulin and phosphatidylserine. Therapies targeting these signals can lead to increased phagocytosis of cancer cells; however, because "don't eat me" signals are markers of "self" on normal cells, treatment can result in negative off-target effects, such as anemia and B-cell depletion. Recent preclinical research has demonstrated the potential of nanocarriers to synergize with prophagocytic therapies, address the off-target effects, improve pharmacokinetics, and codeliver chemotherapeutics. The high surface area-to-volume ratio of nanoparticles paired with preferential size for passive targeting allows for greater accumulation of therapeutic cargo. In addition, nanomaterials hold promise as molecular imaging agents for the detection of phagocytic markers. This mini review highlights the unique capabilities of nanotechnology to expand the application and efficacy of immunotherapy through recently discovered phagocytotic checkpoint therapies.

Preclinical data on co-delivery of temozolomide and talazoparib by fucodain-coated nanoscale metal organic frameworks for colorectal cancer chemoradiation.

Nanoparticle characterization and in vitro data on the effects of combined PARP inhibition and DNA damage by chemoradiation are shown. This data accompanies the research article "Fucoidan-coated nanoparticles target radiation-induced P-selectin to enhance chemoradiotherapy in murine colorectal cancer" (DuRoss et al., 2021) Additional characterization of the physiochemical properties of nanoscale metal organic frameworks (nMOFs) comprised of hafnium and 1,4-dicarboxybenzene (Hf-BDC) loaded with temozolomide (TMZ) and talazoparib (Tal) are presented. Toxicity data of the drug-loaded nMOF coated with fucoidan (TT@Hf-BDC-Fuco) in colorectal cancer cells, CT-26, from alamarBlue-based chemoradiation experiments are shown. Experimental methods for the nanoparticle characterization and cell-based assays of the nMOF formulation are presented.

Fucoidan-coated nanoparticles target radiation-induced P-selectin to enhance chemoradiotherapy in murine colorectal cancer.

Colorectal cancer (CRC) is a leading cause of cancer-related death for both men and women, highlighting the need for new treatment strategies. Advanced disease is often treated with a combination of radiation and cytotoxic agents, such as DNA damage repair inhibitors and DNA damaging agents. To optimize the therapeutic window of these multimodal therapies, advanced nanomaterials have been investigated to deliver sensitizing agents or enhance local radiation dose deposition. In this study, we demonstrate the feasibility of employing an inflammation targeting nanoscale metal-organic framework (nMOF) platform to enhance CRC treatment. This novel formulation incorporates a fucoidan surface coating to preferentially target P-selectin, which is over-expressed or translocated in irradiated tumors. Using this radiation stimulated delivery strategy, a combination PARP inhibitor (talazoparib) and chemotherapeutic (temozolomide) drug-loaded hafnium and 1,4-dicarboxybenzene (Hf-BDC) nMOF was evaluated both in vitro and in vivo. Significantly, these drug-loaded P-selectin targeted nMOFs (TT@Hf-BDC-Fuco) show improved tumoral accumulation over multiple controls and subsequently enhanced therapeutic effects. The integrated radiation and nanoformulation treatment demonstrated improved tumor control (reduced volume, density, and growth rate) and increased survival in a syngeneic CRC mouse model. Overall, the data from this study support the continued investigation of radiation-priming for targeted drug delivery and further consideration of nanomedicine strategies in the clinical management of advanced CRC.

Tb-Doped core-shell-shell nanophosphors for enhanced X-ray induced luminescence and sensitization of radiodynamic therapy.

The development of radiation responsive materials, such as nanoscintillators, enables a variety of exciting new theranostic applications. In particular, the ability of nanophosphors to serve as molecular imaging agents in novel modalities, such as X-ray luminescence computed tomography (XLCT), has gained significant interest recently. Here, we present a radioluminescent nanoplatform consisting of Tb-doped nanophosphors with an unique core/shell/shell (CSS) architecture for improved optical emission under X-ray excitation. Owing to the spatial confinement and separation of luminescent activators, these CSS nanophosphors exhibited bright optical luminescence upon irradiation. In addition to standard physiochemical characterization, these CSS nanophosphors were evaluated for their ability to serve as energy mediators in X-ray stimulated photodynamic therapy, also known as radiodynamic therapy (RDT), through attachment of a photosensitizer, rose bengal (RB). Furthermore, cRGD peptide was used as a model targeting agent against U87 MG glioblastoma cells. In vitro RDT efficacy studies suggested the RGD-CSS-RB in combination with X-ray irradiation could induce enhanced DNA damage and increased cell killing, while the nanoparticles alone are well tolerated. These studies support the utility of CSS nanophosphors and warrants their further development for theranostic applications.

Tomographic molecular imaging of x-ray-excitable nanoparticles.

X-ray luminescence computed tomography (XLCT) is proposed as a new dual molecular/anatomical imaging modality. XLCT is based on the selective excitation and optical detection of x-ray-excitable nanoparticles. As a proof of concept, we built a prototype XLCT system and imaged near-IR-emitting Gd(2)O(2)S:Eu phosphors in various phantoms. Imaging in an optically diffusive medium shows that imaging performance is not affected by optical scatter; furthermore, the linear response of the reconstructed images suggests that XLCT is capable of quantitative imaging.