Host-functionalization of macrin nanoparticles to enable drug loading and control tumor-associated macrophage phenotype.

Macrophages are critical regulators of the tumor microenvironment and often present an immuno-suppressive phenotype, supporting tumor growth and immune evasion. Promoting a robust pro-inflammatory macrophage phenotype has emerged as a therapeutic modality that supports tumor clearance, including through synergy with immune checkpoint therapies. Polyglucose nanoparticles (macrins), which possess high macrophage affinity, are useful vehicles for delivering drugs to macrophages, potentially altering their phenotype. Here, we examine the potential of functionalized macrins, synthesized by crosslinking carboxymethyl dextran with L-lysine, as effective carriers of immuno-stimulatory drugs to tumor-associated macrophages (TAMs). Azide groups incorporated during particle synthesis provided a handle for click-coupling of propargyl-modified ß-cyclodextrin to macrins under mild conditions. Fluorescence-based competitive binding assays revealed the ability of ß-cyclodextrin to non-covalently bind to hydrophobic immuno-stimulatory drug candidates (Keq ~ 103 M-1), enabling drug loading within nanoparticles. Furthermore, transcriptional profiles of macrophages indicated robust pro-inflammatory reprogramming (elevated Nos2 and Il12; suppressed Arg1 and Mrc1 expression levels) for a subset of these immuno-stimulatory agents (UNC2025 and R848). Loading of R848 into the modified macrins improved the drug's effect on primary murine macrophages by three-fold in vitro. Intravital microscopy in IL-12-eYFP reporter mice (24 h post-injection) revealed a two-fold enhancement in mean YFP fluorescence intensity in macrophages targeted with R848-loaded macrins, relative to vehicle controls, validating the desired pro-inflammatory reprogramming of TAMs in vivo by cell-targeted drug delivery. Finally, in an intradermal MC38 tumor model, cyclodextrin-modified macrin NPs loaded with immunostimulatory drugs significantly reduced tumor growth. Therefore, efficient and effective repolarization of tumor-associated macrophages to an M1-like phenotype-via drug-loaded macrins-inhibits tumor growth and may be useful as an adjuvant to existing immune checkpoint therapies.

Injectable Granular Hydrogels Enable Avidity-Controlled Biotherapeutic Delivery.

Protein therapeutics represent a rapidly growing class of pharmaceutical agents that hold great promise for the treatment of various diseases such as cancer and autoimmune dysfunction. Conventional systemic delivery approaches, however, result in off-target drug exposure and a short therapeutic half-life, highlighting the need for more localized and controlled delivery. We have developed an affinity-based protein delivery system that uses guest-host complexation between ß-cyclodextrin (CD, host) and adamantane (Ad, guest) to enable sustained localized biomolecule presentation. Hydrogels were formed by the copolymerization of methacrylated CD and methacrylated dextran. Extrusion fragmentation of bulk hydrogels yielded shear-thinning and self-healing granular hydrogels (particle diameter = 32.4 ± 16.4 µm) suitable for minimally invasive delivery and with a high host capacity for the retention of guest-modified proteins. Bovine serum albumin (BSA) was controllably conjugated to Ad via EDC chemistry without affecting the affinity of the Ad moiety for CD (KD = 12.0 ± 1.81 µM; isothermal titration calorimetry). The avidity of Ad-BSA conjugates was directly tunable through the number of guest groups attached, resulting in a fourfold increase in the complex half-life (t1/2 = 5.07 ± 1.23 h, surface plasmon resonance) that enabled a fivefold reduction in protein release at 28 days. Furthermore, we demonstrated that the conjugation of Ad to immunomodulatory cytokines (IL-4, IL-10, and IFNγ) did not detrimentally affect cytokine bioactivity and enabled their sustained release. Our strategy of avidity-controlled delivery of protein-based therapeutics is a promising approach for the sustained local presentation of protein therapeutics and can be applied to numerous biomedical applications.

Uptake of Cyclodextrin Nanoparticles by Macrophages is Dependent on Particle Size and Receptor-Mediated Interactions.

Physiochemical properties of nanoparticles, such as their size and chemical composition, dictate their interaction with professional phagocytes of the innate immune system. Macrophages, in particular, are key regulators of the immune microenvironment that heavily influence particle biodistribution as a result of their uptake. This attribute enables macrophage-targeted delivery, including for phenotypic modulation. Saccharide-based materials, including polyglucose polymers and nanoparticles, are efficient vehicles for macrophage-targeted delivery. Here, we investigate the influence of particle size on cyclodextrin nanoparticle (CDNP) uptake by macrophages and further examine the receptor-mediated interactions that drive macrophage-targeted delivery. We designed and synthesized CDNPs ranging in size from 25 nm to >100 nm in diameter. Increasing particle size was correlated with greater uptake by macrophages in vitro. Both scavenger receptor A1 and mannose receptor were critical mediators of macrophage-targeted delivery, inhibition of which reduced the extent of uptake. Finally, we investigated the cellular bioavailability of drug-loaded CDNPs using a model anti-inflammatory drug, celastrol, which demonstrated that drug bioactivity is improved by CDNP loading relative to free drug alone. This study thus elucidates the interactions between the polyglucose nanoparticles and macrophages, thereby facilitating their application in macrophage-targeted drug delivery that has applications in the context of tissue injury and repair.

Mesenchymal Stem Cells Delivered Locally to Ischemia-Reperfused Kidneys via Injectable Hyaluronic Acid Hydrogels Decrease Extracellular Matrix Remodeling 1 Month after Injury in Male Mice.

The translation of stem cell therapies has been hindered by low cell survival and retention rates. Injectable hydrogels enable the site-specific delivery of therapeutic cargo, including cells, to overcome these challenges. We hypothesized that delivery of mesenchymal stem cells (MSC) via shear-thinning and injectable hyaluronic acid (HA) hydrogels would mitigate renal damage following ischemia-reperfusion acute kidney injury. Acute kidney injury (AKI) was induced in mice by bilateral or unilateral ischemia-reperfusion kidney injury. Three days later, mice were treated with MSCs either suspended in media injected intravenously via the tail vein, or injected under the capsule of the left kidney, or MSCs suspended in HA injected under the capsule of the left kidney. Serial measurements of serum and urine biomarkers of renal function and injury, as well as transcutaneous glomerular filtration rate (tGFR) were performed. In vivo optical imaging showed that MSCs localized to both kidneys in a sustained manner after bilateral ischemia and remained within the ipsilateral treated kidney after unilateral ischemic AKI. One month after injury, MSC/HA treatment significantly reduced urinary NGAL compared to controls; it did not significantly reduce markers of fibrosis compared to untreated controls. An analysis of kidney proteomes revealed decreased extracellular matrix remodeling and high overlap with sham proteomes in MSC/HA-treated animals. Hydrogel-assisted MSC delivery shows promise as a therapeutic treatment following acute kidney injury.

Novel Therapeutics: Can Hydrogels Work to Treat Kidney Disease?

BACKGROUND: Hydrogels are water-swollen networks that can be made from a variety of natural and synthetic polymers. Numerous chemistries can be utilized to formulate hydrogels that are injectable, enabling facile in situ delivery of therapeutics such as cytokines or cells. SUMMARY: Cells delivered via injectable hydrogels survive injection better than cells injected in saline or media suspension. Several materials have been used to investigate the use of injectable hydrogels to treat animal models of kidney disease. Species studied to date include mice and rats. This review summarizes the various materials, encapsulated therapeutic payloads, and preclinical models of kidney disease employed to investigate hydrogel injection. Transcutaneous measurements of glomerular filtration rate have demonstrated that delivery of hydrogels under the kidney capsule does not impair kidney function. KEY MESSAGES: Studies to date have shown the safety and efficacy of hydrogel therapies to treat kidney disease, and numerous studies have demonstrated that hydrogel therapy alone reduces inflammation and fibrosis.

Control of the post-infarct immune microenvironment through biotherapeutic and biomaterial-based approaches.

Ischemic heart failure (IHF) is a leading cause of morbidity and mortality worldwide, for which heart transplantation remains the only definitive treatment. IHF manifests from myocardial infarction (MI) that initiates tissue remodeling processes, mediated by mechanical changes in the tissue (loss of contractility, softening of the myocardium) that are interdependent with cellular mechanisms (cardiomyocyte death, inflammatory response). The early remodeling phase is characterized by robust inflammation that is necessary for tissue debridement and the initiation of repair processes. While later transition toward an immunoregenerative function is desirable, functional reorientation from an inflammatory to reparatory environment is often lacking, trapping the heart in a chronically inflamed state that perpetuates cardiomyocyte death, ventricular dilatation, excess fibrosis, and progressive IHF. Therapies can redirect the immune microenvironment, including biotherapeutic and biomaterial-based approaches. In this review, we outline these existing approaches, with a particular focus on the immunomodulatory effects of therapeutics (small molecule drugs, biomolecules, and cell or cell-derived products). Cardioprotective strategies, often focusing on immunosuppression, have shown promise in pre-clinical and clinical trials. However, immunoregenerative therapies are emerging that often benefit from exacerbating early inflammation. Biomaterials can be used to enhance these therapies as a result of their intrinsic immunomodulatory properties, parallel mechanisms of action (e.g., mechanical restraint), or by enabling cell or tissue-targeted delivery. We further discuss translatability and the continued progress of technologies and procedures that contribute to the bench-to-bedside development of these critically needed treatments.

T cell-independent eradication of experimental glioma by intravenous TLR7/8-agonist-loaded nanoparticles.

Glioblastoma, the most common and aggressive primary brain tumor type, is considered an immunologically "cold" tumor with sparse infiltration by adaptive immune cells. Immunosuppressive tumor-associated myeloid cells are drivers of tumor progression. Therefore, targeting and reprogramming intratumoral myeloid cells is an appealing therapeutic strategy. Here, we investigate a ß-cyclodextrin nanoparticle (CDNP) formulation encapsulating the Toll-like receptor 7 and 8 (TLR7/8) agonist R848 (CDNP-R848) to reprogram myeloid cells in the glioma microenvironment. We show that intravenous monotherapy with CDNP-R848 induces regression of established syngeneic experimental glioma, resulting in increased survival rates compared with unloaded CDNP controls. Mechanistically, CDNP-R848 treatment reshapes the immunosuppressive tumor microenvironment and orchestrates tumor clearing by pro-inflammatory tumor-associated myeloid cells, independently of T cells and NK cells. Using serial magnetic resonance imaging, we identify a radiomic signature in response to CDNP-R848 treatment and ultrasmall superparamagnetic iron oxide (USPIO) imaging reveals that immunosuppressive macrophage recruitment is reduced by CDNP-R848. In conclusion, CDNP-R848 induces tumor regression in experimental glioma by targeting blood-borne macrophages without requiring adaptive immunity.

Sustained release of drug-loaded nanoparticles from injectable hydrogels enables long-term control of macrophage phenotype.

Injectable hydrogels may be pre-formed through dynamic crosslinks, allowing for injection and subsequent retention in the tissue by shear-thinning and self-healing processes, respectively. These properties enable the site-specific delivery of encapsulated therapeutics; yet, the sustained release of small-molecule drugs and their cell-targeted delivery remains challenging due to their rapid diffusive release and non-specific cellular biodistribution. Herein, we develop an injectable hydrogel system composed of a macrophage-targeted nanoparticle (cyclodextrin nanoparticles, CDNPs) crosslinked by adamantane-modified hyaluronic acid (Ad-HA). The polymer-nanoparticle hydrogel uniquely leverages cyclodextrin's interaction with small molecule drugs to create a spatially discrete drug reservoir and with adamantane to yield dynamic, injectable hydrogels. Through an innovative two-step drug screening approach and examination of 45 immunomodulatory drugs with subsequent in-depth transcriptional profiling of both murine and human macrophages, we identify celastrol as a potent inhibitor of pro-inflammatory (M1-like) behavior that furthermore promotes a reparatory (M2-like) phenotype. Celastrol encapsulation within the polymer-nanoparticle hydrogels permitted shear-thinning injection and sustained release of drug-laden nanoparticles that targeted macrophages to modulate cell behavior for greater than two weeks in vitro. The modular hydrogel system is a promising approach to locally modulate cell-specific phenotype in a range of applications for immunoregenerative medicine.

Measurement of glomerular filtration rate reveals that subcapsular injection of shear-thinning hyaluronic acid hydrogels does not impair kidney function in mice.

The continued development of minimally invasive therapeutic implants, such as injectable hydrogels, necessitates the concurrent advancement of methods to best assess their biocompatibility via functional outcomes in vivo. Biomaterial implants have been studied to treat kidney disease; however, assessment of biocompatibility has been limited to biomarker and histological assessments. Techniques now exist to measure kidney function serially in vivo in murine studies via transcutaneous measurements of glomerular filtration rate (tGFR). In this study, adult male and female wild-type BalbC mice underwent right unilateral nephrectomy. The remaining solitary left kidney was allowed 4 weeks to recover via compensatory hypertrophy, after which subcapsular injection of either saline or shear-thinning hyaluronic acid hydrogel was performed. Serial tGFR measurements before and after treatment were used to assess the effect of hydrogel injection on kidney filtration. Urine and serum biomarkers of kidney function, and kidney histology were also quantified. Hydrogel injection did not affect kidney function, as assessed by tGFR. Results were in agreement with standard metrics of serum and urine biomarkers of injury as well as histological assessment of inflammation. The model developed provides a direct functional assessment of implant compatibility for the treatment of kidney disease and impact on kidney function.

Macrophage-Targeted Therapy Unlocks Antitumoral Cross-talk between IFNγ-Secreting Lymphocytes and IL12-Producing Dendritic Cells.

Macrophages often abound within tumors, express colony-stimulating factor 1 receptor (CSF1R), and are linked to adverse patient survival. Drugs blocking CSF1R signaling have been used to suppress tumor-promoting macrophage responses; however, their mechanisms of action remain incompletely understood. Here, we assessed the lung tumor immune microenvironment in mice treated with BLZ945, a prototypical small-molecule CSF1R inhibitor, using single-cell RNA sequencing and mechanistic validation approaches. We showed that tumor control was not caused by CSF1R+ cell depletion; instead, CSF1R targeting reshaped the CSF1R+ cell landscape, which unlocked cross-talk between antitumoral CSF1R- cells. These cells included IFNγ-producing natural killer and T cells, and an IL12-producing dendritic cell subset, denoted as DC3, which were all necessary for CSF1R inhibitor-mediated lung tumor control. These data indicate that CSF1R targeting can activate a cardinal cross-talk between cells that are not macrophages and that are essential to mediate the effects of T cell-targeted immunotherapies and promote antitumor immunity.See related Spotlight by Burrello and de Visser, p. 4.

Quantification of Cellular Drug Biodistribution Addresses Challenges in Evaluating in vitro and in vivo Encapsulated Drug Delivery.

Nanoencapsulated drug delivery to solid tumors is a promising approach to overcome pharmacokinetic limitations of therapeutic drugs. However, encapsulation leads to complex drug biodistribution and delivery making analysis of delivery efficacy challenging. As proxies, nanocarrier accumulation or total tumor drug uptake in the tumor are used to evaluate delivery. Yet, these measurements fail to assess delivery of active, released drug to the target, and thus it commonly remains unknown if drug-target occupancy has been achieved. Here, we develop an approach to evaluate the delivery of encapsulated drug to the target, where residual drug target vacancy is measured using a fluorescent drug analog. In vitro measurements reveal that burst release governs drug delivery independent of nanoparticle uptake, and highlight limitations of evaluating nanoencapsulated drug delivery in these models. In vivo, however, our approach captures successful nanoencapsulated delivery, finding that tumor stromal cells drive nanoparticle accumulation and mediate drug delivery to adjacent cancer cells. These results, and generalizable approach, provide a critical advance to evaluate delivery of encapsulated drug to the drug target - the central objective of nanotherapeutics.

Therapeutically reprogrammed nutrient signalling enhances nanoparticulate albumin bound drug uptake and efficacy in KRAS-mutant cancer.

Nanoparticulate albumin bound paclitaxel (nab-paclitaxel, nab-PTX) is among the most widely prescribed nanomedicines in clinical use, yet it remains unclear how nanoformulation affects nab-PTX behaviour in the tumour microenvironment. Here, we quantified the biodistribution of the albumin carrier and its chemotherapeutic payload in optically cleared tumours of genetically engineered mouse models, and compared the behaviour of nab-PTX with other clinically relevant nanoparticles. We found that nab-PTX uptake is profoundly and distinctly affected by cancer-cell autonomous RAS signalling, and RAS/RAF/MEK/ERK inhibition blocked its selective delivery and efficacy. In contrast, a targeted screen revealed that IGF1R kinase inhibitors enhance uptake and efficacy of nab-PTX by mimicking glucose deprivation and promoting macropinocytosis via AMPK, a nutrient sensor in cells. This study thus shows how nanoparticulate albumin bound drug efficacy can be therapeutically improved by reprogramming nutrient signalling and enhancing macropinocytosis in cancer cells.

Applications of Macrocyclic Host Molecules in Immune Modulation and Therapeutic Delivery.

The immune system plays a central role in the development and progression of human disease. Modulation of the immune response is therefore a critical therapeutic target that enables us to approach some of the most vexing problems in medicine today such as obesity, cancer, viral infection, and autoimmunity. Methods of manipulating the immune system through therapeutic delivery centralize around two common themes: the local delivery of biomaterials to affect the surrounding tissue or the systemic delivery of soluble material systems, often aided by context-specific cell or tissue targeting strategies. In either case, supramolecular interactions enable control of biomaterial composition, structure, and behavior at the molecular-scale; through rational biomaterial design, the realization of next-generation immunotherapeutics and immunotheranostics is therefore made possible. This brief review highlights methods of harnessing macromolecular interaction for immunotherapeutic applications, with an emphasis on modes of drug delivery.

Polymeric materials for immune engineering: Molecular interaction to biomaterial design.

Biomaterials continue to evolve as complex engineered tools for interactively instructing biological systems, aiding in the understanding and treatment of various disease states through intimate biological interaction. The immune response to polymeric materials is a critical area of study, as it governs the body's response to biomaterial implants, drug delivery vehicles, and even therapeutic drug formulations. Importantly, the development of the immune response to polymeric biomaterials spans length scales - from single molecular interactions to the complex sensing of bulk biophysical properties, all of which coordinate a tissue- and systems-level response. In this review, we specifically discuss a bottom-up approach to designing biomaterials that use molecular-scale interactions to drive immune response to polymers and discuss how these interactions can be leveraged for biomaterial design. STATEMENT OF SIGNIFICANCE: The immune system is an integral controller of (patho)physiological processes, affecting nearly all aspects of human health and disease. Polymeric biomaterials, whether biologically derived or synthetically produced, can potentially alter the behavior of immune cells due to their molecular-scale interaction with individual cells, as well as their interpretation at the bulk scale. This article reviews common mechanisms by which immune cells interact with polymers at the molecular level and discusses how these interactions are being leveraged to produce the next generation of biocompatible and immunomodulatory materials.

Tunable Blood Shunt for Neonates With Complex Congenital Heart Defects.

Despite advancements in procedures and patient care, mortality rates for neonatal recipients of the Norwood procedure, a palliation for single ventricle congenital malformations, remain high due to the use of a fixed-diameter blood shunt. In this study, a new geometrically tunable blood shunt was investigated to address limitations of the current treatment paradigm (e.g., Modified Blalock-Taussig Shunt) by allowing for controlled modulation of blood flow through the shunt to accommodate physiological changes due to the patient's growth. First, mathematical and computational cardiovascular models were established to investigate the hemodynamic requirements of growing neonatal patients with shunts and to inform design criteria for shunt diameter changes. Then, two stages of prototyping were performed to design, build and test responsive hydrogel systems that facilitate tuning of the shunt diameter by adjusting the hydrogel's degree of crosslinking. We examined two mechanisms to drive crosslinking: infusion of chemical crosslinking agents and near-UV photoinitiation. The growth model showed that 15-18% increases in shunt diameter were required to accommodate growing patients' increasing blood flow; similarly, the computational models demonstrated that blood flow magnitudes were in agreement with previous reports. These target levels of diameter increases were achieved experimentally with model hydrogel systems. We also verified that the photocrosslinkable hydrogel, composed of methacrylated dextran, was contact-nonhemolytic. These results demonstrate proof-of-concept feasibility and reflect the first steps in the development of this novel blood shunt. A tunable shunt design offers a new methodology to rebalance blood flow in this vulnerable patient population during growth and development.

How hydrogel inclusions modulate the local mechanical response in early and fully formed post-infarcted myocardium.

Expansion of myocardium after myocardial infarction (MI) has long been identified as the primary mechanism that drives adverse left ventricular (LV) remodeling towards heart failure and death. Direct injection of hydrogels into the myocardium to mechanically constrain the infarct has demonstrated promise in limiting its remodeling and expansion. Despite early successes, there remain open questions in the determination of optimal hydrogel therapies, key application characteristics for which include injected polymer volume, stiffness, and spatial placement. Addressing these questions is complicated by the substantial variations in infarct type and extent, as well as limited understanding of the underlying mechanisms. Herein, we present an investigation on how hydrogel inclusions affect the effective tissue-level stiffness and strain fields in myocardium using full three-dimensional (3D) finite element simulations at early and late post-MI time points. We calibrated our simulations to triaxial mechanical and structural measurements of cuboidal LV myocardial specimens of post-infarcted myocardium, 0 and 4 weeks post-MI, injected with a dual-crosslinking hyaluronic acid-based hydrogel. Simulations included multiple deformation modes that spanned the anticipated physiological range in order to assess the effects of variations in inclusion size, location, and modulus on tissue-level myocardial mechanics. We observed significant local stiffening in the hydrogel-injected specimens that was highly dependent on the volume and mechanical properties of the injected hydrogel. Simulations revealed that the primary effect of the injections under physiological loading was a reduction in myocardial strain. This result suggests that hydrogel injections reduce infarct expansion by limiting the peak strains over the cardiac cycle. Overall, our study indicated that modulation of local effective tissue stiffness and corresponding strain reduction are governed by the volume and stiffness of the hydrogel, but relatively insensitive to its transmural placement. These findings provide important insights into mechanisms for ameliorating post-MI remodeling, as well as guidance for the future design of post-MI therapies.

Myeloid Cell-Targeted Nanocarriers Efficiently Inhibit Cellular Inhibitor of Apoptosis for Cancer Immunotherapy.

Immune-checkpoint blockers can promote sustained clinical responses in a subset of cancer patients. Recent research has shown that a subpopulation of tumor-infiltrating dendritic cells functions as gatekeepers, sensitizing tumors to anti-PD-1 treatment via production of interleukin-12 (IL-12). Hypothesizing that myeloid cell-targeted nanomaterials could be used to deliver small-molecule IL-12 inducers, we performed high-content image-based screening to identify the most efficacious small-molecule compounds. Using one lead candidate, LCL161, we created a myeloid-targeted nanoformulation that induced IL-12 production in intratumoral myeloid cells in vivo, slowed tumor growth as a monotherapy, and had no significant systemic toxicity. These results pave the way for developing combination immunotherapeutics by harnessing IL-12 production for immunostimulation.

Injectable Shear-Thinning Hydrogels Prevent Ischemic Mitral Regurgitation and Normalize Ventricular Flow Dynamics.

Injectable hydrogels are known to attenuate left-ventricular (LV) remodeling following myocardial infarction (MI), dependent on material mechanical properties. The effect of hydrogel injection on ischemic mitral regurgitation (IMR) resultant from LV remodeling remains relatively unexplored. This study uses multiple imaging methods to evaluate the efficacy of injectable hydrogels with tunable modulus to prevent post-MI development of IMR. Posterolateral MI was induced in 20 sheep with subsequent epicardial injection of saline (control (MI); n = 7), soft hydrogel (guest-host crosslinking, modulus <1 kPa, n = 7), or stiff hydrogel (dual-crosslinking, modulus = 41.4 ± 4.3 kPa, n = 6) within the infarct region and 8-week follow-up. IMR and valve geometry were assessed by echocardiography. LV geometry (long-axis dimension, posterior chordae length) and ventricular flow dynamics were assessed by magnetic resonance imaging. IMR developed in MI controls at 8 weeks and was attenuated with hydrogel treatment (IMR grade for MI: 1.86 ± 0.69; guest-host crosslinking: 1.29 ± 1.11; dual-crosslinking: 0.50 ± 0.55, P = 0.02 vs MI). Tethering of the posterior leaflet increased in MI controls, but not with stiff hydrogel treatment. Across cohorts, IMR was correlated with changes in the long-axis dimension (Spearman R = 0.77) and posterior chordae length (Spearman R = 0.64). Intraventricular flow dynamics were highly disturbed in MI controls, but stiff hydrogel treatment normalized flow patterns and reduced the prevalence of large (≥2+ MR, >5 mL) regurgitant volumes. Injectable hydrogels attenuated subvalvular remodeling and leaflet tethering, preventing IMR development and normalizing LV flow dynamics. Hydrogels with a supraphysiological modulus yielded best outcomes.

Development of Adamantane-Conjugated TLR7/8 Agonists for Supramolecular Delivery and Cancer Immunotherapy.

Tumor-associated macrophages (TAMs) are often abundant in solid cancers, assuming an immunosuppressive (M2-like) phenotype which supports tumor growth and immune escape. Recent methods have focused on identification of means (e.g., drugs, nanomaterials) that polarize TAMs to a tumor suppressive (M1-like) phenotype; however, reducing the systemic side effects of these therapies and enabling their delivery to TAMs has remained a challenge. Methods: Here, we develop R848-Ad, an adamantane-modified derivative of the toll-like receptor (TLR) 7/8 agonist resiquimod (R848) through iterative drug screening against reporter cell lines. The adamantane undergoes guest-host interaction with cyclodextrin nanoparticles (CDNPs), enabling drug loading under aqueous conditions and TAM-targeted drug delivery. Therapeutic efficacy and systemic side effects were examined in a murine MC38 cancer model. Results: R848-Ad retained macrophage polarizing activity through agonization of TLR7/8, and the adamantane moiety improved drug affinity for the CDNP. In preclinical studies, nanoformulated R848-Ad resulted in a drastic reduction in measurable systemic effects (loss of body weight) relative to similarly formulated R848 alone while arresting tumor growth. Conclusions: The findings demonstrate the ability of strong nanoparticle-drug interactions to limit systemic toxicity of TLR agonists while simultaneously maintaining therapeutic efficacy.