Precise modulation and use of reactive oxygen species for immunotherapy.

Reactive oxygen species (ROS) play an important role in regulating the immune system by affecting pathogens, cancer cells, and immune cells. Recent advances in biomaterials have leveraged this mechanism to precisely modulate ROS levels in target tissues for improving the effectiveness of immunotherapies in infectious diseases, cancer, and autoimmune diseases. Moreover, ROS-responsive biomaterials can trigger the release of immunotherapeutics and provide tunable release kinetics, which can further boost their efficacy. This review will discuss the latest biomaterial-based approaches for both precise modulation of ROS levels and using ROS as a stimulus to control the release kinetics of immunotherapeutics. Finally, we will discuss the existing challenges and potential solutions for clinical translation of ROS-modulating and ROS-responsive approaches for immunotherapy, and provide an outlook for future research.

A Virus-Inspired Inhalable Liponanogel Induces Potent Antitumor Immunity and Regression in Metastatic Lung Tumors.

Pulmonary delivery of immunostimulatory agents such as poly(I:C) to activate double-stranded RNA sensors MDA5 and RIG-I within lung-resident antigen-presenting cells is a potential strategy to enhance antitumor immunity by promoting type I interferon secretion. However, following pulmonary delivery, poly(I:C) suffers from rapid degradation and poor endosomal escape, thus limiting its potency. Inspired by the structure of a virus that utilizes internal viral proteins to tune the loading and cytosolic delivery of viral nucleic acids, we developed a liponanogel (LNG)-based platform to overcome the delivery challenges of poly(I:C). The LNG consisted of an anionic polymer hyaluronic acid-based nanogel core coated by a lipid shell, which served as a protective layer to stabilize the nanogel core in the lungs. The nanogel core was protonated within acidic endosomes to enhance the endosomal membrane permeability and cytosolic delivery of poly(I:C). After pulmonary delivery, LNG-poly(I:C) induced 13.7-fold more IFNß than poly(I:C) alone and 2-fold more than poly(I:C) loaded in the state-of-art lipid nanoparticles (LNP-poly(I:C)). Moreover, LNG-poly(I:C) induced more potent CD8+ T cell immunity and stronger therapeutic effects than LNP-poly(I:C). The combination of LNG-poly(I:C) and PD-L1 targeting led to regression of established lung metastases. Due to the ease of manufacturing and the high biocompatibility of LNG, pulmonary delivery of LNG may be broadly applicable to the treatment of different lung tumors and may spur the development of innovative strategies for cancer immunotherapy.

Ultrasound-responsive low-dose doxorubicin liposomes trigger mitochondrial DNA release and activate cGAS-STING-mediated antitumour immunity.

DNA derived from chemotherapeutics-killed tumor cells is one of the most important damage-associated molecular patterns that can activate the cGAS-STING (cyclic GMP-AMP synthase-stimulator of interferon genes) pathway in antigen-presenting cells (APCs) and promote antitumor immunity. However, conventional chemotherapy displays limited tumor cell killing and ineffective transfer of stable tumor DNA to APCs. Here we show that liposomes loaded with an optimized ratio of indocyanine green and doxorubicin, denoted as LID, efficiently generate reactive oxygen species upon exposure to ultrasound. LID plus ultrasound enhance the nuclear delivery of doxorubicin, induce tumor mitochondrial DNA oxidation, and promote oxidized tumor mitochondrial DNA transfer to APCs for effective activation of cGAS-STING signaling. Depleting tumor mitochondrial DNA or knocking out STING in APCs compromises the activation of APCs. Furthermore, systemic injection of LID plus ultrasound over the tumor lead to targeted cytotoxicity and STING activation, eliciting potent antitumor T cell immunity, which upon the combination with immune checkpoint blockade leads to regression of bilateral MC38, CT26, and orthotopic 4T1 tumors in female mice. Our study sheds light on the importance of oxidized tumor mitochondrial DNA in STING-mediated antitumor immunity and may inspire the development of more effective strategies for cancer immunotherapy.

Clathrin light chain-conjugated drug delivery for cancer.

Targeted drug delivery systems hold the remarkable potential to improve the therapeutic index of anticancer medications markedly. Here, we report a targeted delivery platform for cancer treatment using clathrin light chain (CLC)-conjugated drugs. We conjugated CLC to paclitaxel (PTX) through a glutaric anhydride at high efficiency. Labeled CLCs localized to 4T1 tumors implanted in mice, and conjugation of PTX to CLC enhanced its delivery to these tumors. Treatment of three different mouse models of cancer-melanoma, breast cancer, and lung cancer-with CLC-PTX resulted in significant growth inhibition of both the primary tumor and metastatic lesions, as compared to treatment with free PTX. CLC-PTX treatment caused a marked increase in apoptosis of tumor cells and reduction of tumor angiogenesis. Our data suggested HSP70 as a binding partner for CLC. Our study demonstrates that CLC-based drug-conjugates constitute a novel drug delivery platform that can augment the effects of chemotherapeutics in treating a variety of cancers. Moreover, conjugation of therapeutics with CLC may be used as means by which drugs are delivered specifically to primary tumors and metastatic lesions, thereby prolonging the survival of cancer patients.

Systematic evaluation of the effect of different apolipoprotein A-I mimetic peptides on the performance of synthetic high-density lipoproteins in vitro and in vivo.

Synthetic high-density lipoproteins nanomedicine (sHDL) composed of apolipoprotein A-I (ApoA-I) mimetic peptides and lipids have shown very promising results for the treatment of various cardiovascular diseases. Numerous efforts have also been made to design different ApoA-I mimetic peptides to improve the potency of sHDL, especially the efficiency of reverse cholesterol transport. However, the way in which ApoA-I mimetic peptides affect the properties of sHDL, including stability, cholesterol efflux, cholesterol esterification, elimination in vivo, and the relationship of these properties, is still poorly understood. Revealing the effect of these factors on the potency of sHDL is important for the design of better ApoA-I mimetic peptides. In this study, three widely used ApoA-I mimetic peptides with different sequences, lengths, LCAT activation and lipid binding affinities were used for the preparation of sHDL and were evaluated in terms of physical/chemical properties, cholesterol efflux, cholesterol esterification, remodeling, and pharmacokinetics/pharmacodynamics. Our results showed that ApoA-I mimetic peptides with the highest cholesterol efflux and cholesterol esterification in vitro did not exhibit the highest cholesterol mobilization in vivo. Further analysis indicated that other factors, such as pharmacokinetics and remodeling of sHDL, need to be considered in order to predict the efficiency of cholesterol mobilization in vivo. Thus, our study highlights the importance of using the overall performance, rather than in vitro results alone, as the blueprint for the design and optimization of ApoA-I mimetic peptides.

A cell-based drug delivery platform for treating central nervous system inflammation.

Mesenchymal stem cells (MSCs) are promising candidates for the development of cell-based drug delivery systems for autoimmune inflammatory diseases, such as multiple sclerosis (MS). Here, we investigated the effect of Ro-31-8425, an ATP-competitive kinase inhibitor, on the therapeutic properties of MSCs. Upon a simple pretreatment procedure, MSCs spontaneously took up and then gradually released significant amounts of Ro-31-8425. Ro-31-8425 (free or released by MSCs) suppressed the proliferation of CD4+ T cells in vitro following polyclonal and antigen-specific stimulation. Systemic administration of Ro-31-8425-loaded MSCs ameliorated the clinical course of experimental autoimmune encephalomyelitis (EAE), a murine model of MS, displaying a stronger suppressive effect on EAE than control MSCs or free Ro-31-8425. Ro-31-8425-MSC administration resulted in sustained levels of Ro-31-8425 in the serum of EAE mice, modulating immune cell trafficking and the autoimmune response during EAE. Collectively, these results identify MSC-based drug delivery as a potential therapeutic strategy for the treatment of autoimmune diseases. KEY MESSAGES: MSCs can spontaneously take up the ATP-competitive kinase inhibitor Ro-31-8425. Ro-31-8425-loaded MSCs gradually release Ro-31-8425 and exhibit sustained suppression of T cells. Ro-31-8425-loaded MSCs have more sustained serum levels of Ro-31-8425 than free Ro-31-8425. Ro-31-8425-loaded MSCs are more effective than MSCs and free Ro-31-8425 for EAE therapy.

Synthetic high-density lipoproteins delivering liver X receptor agonist prevent atherogenesis by enhancing reverse cholesterol transport.

Liver X nuclear receptor (LXR) agonists are promising anti-atherosclerotic agents that increase the expression of cholesterol transporters on atheroma macrophages leading to increased efflux of cholesterol to endogenous high-density lipoprotein (HDL) acceptors. HDL subsequently delivers effluxed cholesterol to the liver by the process of reverse cholesterol transport, resulting in reduction of atherosclerotic plaques. However, LXR agonists administration triggers undesirable liver steatosis and hypertriglyceridemia due to increased fatty acid and sterol synthesis. LXR-induced liver toxicity, poor drug aqueous solubility and low levels of endogenous HDL acceptors in target patient populations limit the clinical translation of LXR agonists. Here, we propose a dual-antiatherogenic strategy for administration of the LXR agonist, T0901317 (T1317), by encapsulating in synthetic HDL (sHDL) nanoparticles. sHDL had been clinically proven to serve as cholesterol acceptors, resulting in plaque reduction in atherosclerosis patients. In addition, the hydrophobic core and endogenous atheroma-targeting ability of sHDL allow for encapsulation of water-insoluble drugs and their subsequent delivery to atheroma. Several compositions of sHDL were tested to optimize both T1317 encapsulation efficiency and ability of T1317-sHDL to efflux cholesterol. Optimized T1317-sHDL exhibited more efficient cholesterol efflux from macrophages and enhanced atheroma-targeting relative to free drug. Most importantly, in an apolipoprotein E deficient (ApoE-/-) atherosclerosis progression murine model, T1317-sHDL showed superior inhibition of atherogenesis and reduced hypertriglyceridemia side effects in comparison to the free drug and blank sHDL. The T1317-sHDL pharmacological efficacy was observed at doses lower than those previously described for LXR agents, which may have additional safety benefits. In addition, the established clinical manufacturing, safety and efficacy of blank sHDL nanoparticles used in this study could facilitate future clinical translation of LXR-loaded sHDLs.

RalA exerts an inhibitory effect on IL-1ß/IL-18 secretion by blocking NLRP3 inflammasome activation in levornidazole-treated human THP-1 macrophages.

The NLRP3 inflammasome is an important mediator of inflammatory responses and its regulation is an active area of research. RalA is a Ras-like GTPase, which play pivotal roles in the biology of cells. So far, there have been very few studies on RalA regulating inflammatory responses. Bioinformatics analysis predicted that RalA might participate in the regulatory network of NLRP3 inflammasome, which has been confirmed in THP-1 macrophages. After virtual screening of compounds, it was found that levonidazole selected from our virtual small molecule compound library has the potential to bind to RalA. Of note, the interaction of RalA/levornidazole was verified by Surface Plasmon Resonance-Biacore T200, LC/MS analysis and Western blotting analysis. Molecular dynamics simulations revealed that the conformational changes of RalA might be regulated by levornidazole. Additionally, IL-1ß/IL-18 secretion from ATP + LPS stimulated THP-1-derived macrophages was RalA-dependently suppressed by levornidazole, suggesting that RalA might have an inhibitory effect on NLRP3 inflammasome activation. The results of co-immunoprecipitation and RalA depletion experiments showed that levornidazole could induce RalA to block the assembly of NLRP3/ASC/pro-caspase-1 complex, thereby reducing the levels of cleaved-caspase-1 and IL-1ß/IL-18 secretion. Our study has suggested an anti-inflammatory function of RalA and identified its targeting chemical compound. Overall, this study clarifies a novel pharmacological mechanism by which RalA/levornidazole inhibits NLRP3 inflammasome activation and IL-1ß/IL-18 secretion.

Shattering barriers toward clinically meaningful MSC therapies.

More than 1050 clinical trials are registered at FDA.gov that explore multipotent mesenchymal stromal cells (MSCs) for nearly every clinical application imaginable, including neurodegenerative and cardiac disorders, perianal fistulas, graft-versus-host disease, COVID-19, and cancer. Several companies have or are in the process of commercializing MSC-based therapies. However, most of the clinical-stage MSC therapies have been unable to meet primary efficacy end points. The innate therapeutic functions of MSCs administered to humans are not as robust as demonstrated in preclinical studies, and in general, the translation of cell-based therapy is impaired by a myriad of steps that introduce heterogeneity. In this review, we discuss the major clinical challenges with MSC therapies, the details of these challenges, and the potential bioengineering approaches that leverage the unique biology of MSCs to overcome the challenges and achieve more potent and versatile therapies.

Robust anti-tumor T cell response with efficient intratumoral infiltration by nanodisc cancer immunotherapy.

Potent anti-tumor T cell response and efficient intratumoral T cell infiltration are the major challenges for therapeutic cancer vaccines. To address these issues, a nano-vaccine system has been designed to promote anti-tumor T cell responses, and intratumoral infiltration was examined in various murine tumor models. Subcutaneous vaccination with nanodiscs carrying human papillomavirus (HPV)-16 E7 antigen elicits as high as ~32% E7-specific CD8 α + T cell responses in circulation, representing a 29-fold improvement over the soluble peptide vaccination. Importantly, nanodisc vaccination also promotes robust intratumoral T cell infiltration and eliminates HPV16 E6/E7-expressing TC-1 tumors at mucosal sites, including lungs, inner lip, and intravaginal tissues. In a benchmark study with a live Listeria vaccine combined with anti-PD-1 IgG, nanodiscs plus anti-PD-1 immune checkpoint blockade elicits comparable levels of T cell responses with anti-tumor efficacy. Furthermore, compared with Complete Freund's Adjuvant combined with tetanus toxoid, nanodisc vaccination in HLA-A02 mice generates >200-fold stronger IFN-γ+ T cell responses against a neoantigen from an HLA-A02 melanoma patient. Overall, these results show that the nanodisc system is a promising cancer vaccine platform for inducing anti-tumor T cell responses.

Mimetic sHDL nanoparticles: A novel drug-delivery strategy to target triple-negative breast cancer.

BACKGROUND: Withanolides are naturally derived heat shock protein 90 inhibitors that are potent in preclinical models of triple negative breast cancers. Conjugation to synthetic high-density lipoprotein nanoparticles improves solubility and targets delivery to the scavenger receptor B1. Triple negative breast cancers highly overexpress the scavenger receptor B1, and we hypothesize that encapsulation of the novel withalongolide A 4,19,27-triacetate by synthetic high-density lipoprotein will have enhanced efficacy against triple negative breast cancers in vivo. METHODS: Validated human triple negative breast cancer cell lines were evaluated for the scavenger receptor B1 expression by quantitative polymerase chain reaction and Western blot. Withalongolide A 4,19,27-triacetate inhibitory concentration50 values were obtained using CellTiter-Glo assays (Promega, Madison, WI, USA). The scavenger receptor B1-mediated drug uptake was evaluated in vitro under fluorescence microscopy and in vivo with IVIS imaging of mouse xenografts (MD-MBA-468LN). To evaluate drug efficacy, mice were treated with synthetic high-density lipoprotein alone, withalongolide A 4,19,27-triacetate alone, withalongolide A 4,19,27-triacetate synthetic high-density lipoprotein, and chemotherapy or Prussian blue stain (control). RESULTS: Triple negative breast cancer cell lines had greater scavenger receptor B1 expression by quantitative polymerase chain reaction and Western blot versus controls. Fluorescent-labeled synthetic high-density lipoprotein uptake was scavenger receptor B1-mediated in vitro, and in vivo tumor uptake using IVIS imaging demonstrated significantly increased tumor radiant efficiency versus control. Inhibitory concentration50 for withalongolide A 4,19,27-triacetate-treated cells with or without synthetic high-density lipoprotein encapsulation were 70-fold to 200-fold more potent than synthetic high-density lipoprotein alone. In triple negative breast cancer mouse xenografts, treatment with synthetic high-density lipoprotein withalongolide A 4,19,27-triacetate resulted in a 54% decrease in tumor volume compared with the control or with synthetic high-density lipoprotein alone. CONCLUSION: The synthetic high-density lipoprotein withalongolide A 4,19,27-triacetate nanoconjugates are potent against triple negative breast cancers and show improved scavenger receptor B1-mediated targeting. Treatment with synthetic high-density lipoprotein-encapsulated withalongolide A 4,19,27-triacetate is able to significantly decrease the growth of tumor in mice compared with the control and has better efficacy than the current standard of care, warranting further evaluation as a novel therapeutic agent.

High-Density Lipoprotein-Mimicking Nanodiscs for Chemo-immunotherapy against Glioblastoma Multiforme.

Glioblastoma multiforme (GBM) is an aggressive primary brain tumor, for which there is no cure. Treatment effectiveness for GBM has been limited due to tumor heterogeneity, an immunosuppressive tumor microenvironment (TME), and the presence of the blood-brain barrier, which hampers the transport of chemotherapeutic compounds to the central nervous system (CNS). High-density lipoprotein (HDL)-mimicking nanodiscs hold considerable promise to achieve delivery of bioactive compounds into tumors. Herein, we tested the ability of synthetic HDL nanodiscs to deliver chemotherapeutic agents to the GBM microenvironment and elicit tumor regression. To this end, we developed chemo-immunotherapy delivery vehicles based on sHDL nanodiscs loaded with CpG, a Toll-like receptor 9 (TLR9) agonist, together with docetaxel (DTX), a chemotherapeutic agent, for targeting GBM. Our data show that delivery of DTX-sHDL-CpG nanodiscs into the tumor mass elicited tumor regression and antitumor CD8+ T cell responses in the brain TME. We did not observe any overt off-target side effects. Furthermore, the combination of DTX-sHDL-CpG treatment with radiation (IR), which is the standard of care for GBM, resulted in tumor regression and long-term survival in 80% of GBM-bearing animals. Mice remained tumor-free upon tumor cell rechallenge in the contralateral hemisphere, indicating the development of anti-GBM immunological memory. Collectively, these data indicate that sHDL nanodiscs constitute an effective drug delivery platform for the treatment of GBM, resulting in tumor regression, long-term survival, and immunological memory when used in combination with IR. The proposed delivery platform has significant potential for clinical translation.

PEGylated tumor cell membrane vesicles as a new vaccine platform for cancer immunotherapy.

Despite the promise and advantages of autologous cancer cell vaccination, it remains challenging to induce potent anti-tumor immune responses with traditional immunization strategies with whole tumor cell lysate. In this study, we sought to develop a simple and effective approach for therapeutic vaccination with autologous whole tumor cell lysate. Endogenous cell membranes harvested from cancer cells were formed into PEGylated nano-vesicles (PEG-NPs). PEG-NPs exhibited good serum stability in vitro and draining efficiency to local lymph nodes upon subcutaneous administration in vivo. Vaccination with PEG-NPs synthesized from murine melanoma cells elicited 3.7-fold greater antigen-specific cytotoxic CD8+ T lymphocyte responses, compared with standard vaccination with freeze-thawed lysate in tumor-bearing mice. Importantly, in combination with anti-programmed death-1 (αPD-1) IgG immunotherapy, PEG-NP vaccination induced 4.2-fold higher frequency of antigen-specific T cell responses (P < 0.0001) and mediated complete tumor regression in 63% of tumor-bearing animals (P < 0.01), compared with FT lysate + αPD-1 treatment that exhibited only 13% response rate. In addition, PEG-NPs + αPD-1 IgG combination immunotherapy protected all survivors against a subsequent tumor cell re-challenge. These results demonstrate a general strategy for eliciting anti-tumor immunity using endogenous cancer cell membranes formulated into stable vaccine nanoparticles.

Synthetic high-density lipoprotein nanoconjugate targets neuroblastoma stem cells, blocking migration and self-renewal.

BACKGROUND: Pathways critical for neuroblastoma cancer stem cell function are targeted by 4,19,27-triacetyl withalongolide A (WGA-TA). Because neuroblastoma cells and their cancer stem cells highly overexpress the scavenger receptor class B type 1 receptor that binds to synthetic high-density lipoprotein, we hypothesized that a novel mimetic synthetic high-density lipoprotein nanoparticle would be an ideal carrier for the delivery of 4,19,27-triacetyl withalongolide to neuroblastoma and neuroblastoma cancer stem cells. METHODS: Expression of scavenger receptor class B type 1 in validated human neuroblastoma cells was evaluated by quantitative polymerase chain reaction (qPCR) and Western blot. In vitro cellular uptake of synthetic high-density lipoprotein nanoparticles was observed with a fluorescence microscope. In vivo biodistribution of synthetic high-density lipoprotein nanoparticles was investigated with IVIS imaging. Self-renewal and migration/invasion were assessed by sphere formation and Boyden chamber assays, respectively. Viability was analyzed by CellTiter-Glo assay. Cancer stem cell markers were evaluated by flow cytometry. RESULTS: qPCR and Western blot analysis revealed a higher level of scavenger receptor class B type 1 expression and drug uptake in N-myc amplified neuroblastoma cells. In vitro uptake of synthetic high-density lipoprotein was almost completely blocked by excess synthetic high-density lipoprotein. The synthetic high-density lipoprotein nanoparticles mainly accumulated in the tumor and liver, but not in other organs. Synthetic HDL-4,19,27-triacetyl withalongolide showed a 1,000-fold higher potency than the carrier (synthetic high-density lipoprotein) alone (P < .01) to kill neuroblastoma cells. Additionally, a dose-dependent decrease in sphere formation, invasion, migration, and cancer stem cell markers was observed after treatment of neuroblastoma cells with synthetic high-density lipoprotein-4,19,27-triacetyl withalongolide A. CONCLUSION: Synthetic high-density lipoprotein is a promising platform to improve the delivery of anticancer drug 4,19,27-triacetyl withalongolide A to neuroblastomas and neuroblastoma cancer stem cells through SR-B1 targeting in vitro and in vivo.

Elimination of established tumors with nanodisc-based combination chemoimmunotherapy.

Although immune checkpoint blockade has shown initial success for various cancers, only a small subset of patients benefits from this therapy. Some chemotherapeutic drugs have been reported to induce antitumor T cell responses, prompting a number of clinical trials on combination chemoimmunotherapy. However, how to achieve potent immune activation with traditional chemotherapeutics in a manner that is safe, effective, and compatible with immunotherapy remains unclear. We show that high-density lipoprotein-mimicking nanodiscs loaded with doxorubicin (DOX), a widely used chemotherapeutic agent, can potentiate immune checkpoint blockade in murine tumor models. Delivery of DOX via nanodiscs triggered immunogenic cell death of cancer cells and exerted antitumor efficacy without any overt off-target side effects. "Priming" tumors with DOX-carrying nanodiscs elicited robust antitumor CD8+ T cell responses while broadening their epitope recognition to tumor-associated antigens, neoantigens, and intact whole tumor cells. Combination chemoimmunotherapy with nanodiscs plus anti-programmed death 1 therapy induced complete regression of established CT26 and MC38 colon carcinoma tumors in 80 to 88% of animals and protected survivors against tumor recurrence. Our work provides a new, generalizable framework for using nanoparticle-based chemotherapy to initiate antitumor immunity and sensitize tumors to immune checkpoint blockade.

Dual TLR agonist nanodiscs as a strong adjuvant system for vaccines and immunotherapy.

Recent studies have shown that certain combinations of Toll-like receptor (TLR) agonists can induce synergistic immune activation. However, it remains challenging to achieve such robust responses in vivo in a manner that is effective, facile, and amenable for clinical translation. Here, we show that MPLA, a TLR4 agonist, and CpG, a TLR9 agonist, can be efficiently co-loaded into synthetic high-density lipoprotein nanodiscs, forming a potent adjuvant system (ND-MPLA/CpG) that can be readily combined with a variety of subunit antigens, including proteins and peptides. ND-MPLA/CpG significantly enhanced activation of dendritic cells, compared with free dual adjuvants or nanodiscs delivering a single TLR agonist. Importantly, mice immunized with physical mixtures of protein antigens ND-MPLA/CpG generated strong humoral responses, including induction of IgG responses against protein convertase subtilisin/kexin 9 (PCSK9), leading to 17-30% reduction of the total plasma cholesterol levels. Moreover, ND-MPLA/CpG exerted strong anti-tumor efficacy in multiple murine tumor models. Compared with free adjuvants, ND-MPLA/CpG admixed with ovalbumin markedly improved antigen-specific CD8+ T cell responses by 8-fold and promoted regression of B16F10-OVA melanoma (P < 0.0001). Furthermore, ND-MPLA/CpG admixed with E7 peptide antigen elicited ~20% E7-specific CD8+ T cell responses and achieved complete regression of established TC-1 tumors in all treated animals. Taken together, our work highlights the simplicity, versatility, and potency of dual TLR agonist nanodiscs for applications in vaccines and cancer immunotherapy.

Chemo-photothermal therapy combination elicits anti-tumor immunity against advanced metastatic cancer.

Photothermal therapy (PTT) is a promising cancer treatment modality, but PTT generally requires direct access to the source of light irradiation, thus precluding its utility against disseminated, metastatic tumors. Here, we demonstrate that PTT combined with chemotherapy can trigger potent anti-tumor immunity against disseminated tumors. Specifically, we have developed polydopamine-coated spiky gold nanoparticles as a new photothermal agent with extensive photothermal stability and efficiency. Strikingly, a single round of PTT combined with a sub-therapeutic dose of doxorubicin can elicit robust anti-tumor immune responses and eliminate local as well as untreated, distant tumors in >85% of animals bearing CT26 colon carcinoma. We also demonstrate their therapeutic efficacy against TC-1 submucosa-lung metastasis, a highly aggressive model for advanced head and neck squamous cell carcinoma (HNSCC). Our study sheds new light on a previously unrecognized, immunological facet of chemo-photothermal therapy and may lead to new therapeutic strategies against advanced cancer.

Subcutaneous Nanodisc Vaccination with Neoantigens for Combination Cancer Immunotherapy.

While cancer immunotherapy provides new exciting treatment options for patients, there is an urgent need for new strategies that can synergize with immune checkpoint blockers and boost the patient response rates. We have developed a personalized vaccine nanodisc platform based on synthetic high-density lipoproteins for co-delivery of immunostimulatory agents and tumor antigens, including tumor-specific neoantigens. Here we examined the route of delivery, safety profiles, and therapeutic efficacy of nanodisc vaccination against established tumors. We report that nanodiscs administered via the subcutaneous (SC) or intramuscular (IM) routes were well tolerated in mice without any signs of toxicity. The SC route significantly enhanced nanoparticle delivery to draining lymph nodes, improved nanodisc uptake by antigen-presenting cells, and generated 7-fold higher frequency of neoantigen-specific T cells, compared with the IM route. Importantly, when mice bearing advanced B16F10 melanoma tumors were treated with nanodiscs plus anti-PD-1 and anti-CTLA-4 IgG therapy, the combination immunotherapy exerted potent antitumor efficacy, leading to eradication of established tumors in ∼60% of animals. These results demonstrate nanodiscs customized with patient-specific tumor neoepitopes as a safe and powerful vaccine platform for immunotherapy against advanced cancer.

High-density lipoprotein-mimicking nanodiscs carrying peptide for enhanced therapeutic angiogenesis in diabetic hindlimb ischemia.

Therapeutic strategies using endogenous stem cell mobilizer can provide effective cell-free therapy for addressing various ischemic diseases. In particular, substance P (SP) exhibited therapeutic regeneration by facilitating mobilization of endogenous stem cells from bone marrow to the injured sites. However, its therapeutic effect has been limited due to short half-life and rapid degradation of administered SP peptides in vivo. Here we sought to develop high-density lipoprotein (HDL)-mimicking nanodiscs conjugated with SP (HDL-SP) in order to increase the in vivo half-life, bone marrow targeting, and therapeutic efficacy of SP for the treatment of diabetic peripheral ischemia. Conjugation of SP onto HDL nanodisc led to remarkable ∼3215- and ∼1060-fold increase in the ex vivo and in vivo half-lives of SP, respectively. Accordingly, HDL-SP nanodiscs improved retention of SP in bone marrow after systemic administration, leading to efficient mobilization of stem cells from bone marrow into blood circulation and reduction of systemic inflammation. Consequently, nanodisc based SP peptide delivery promoted blood vessel formation, blood perfusion recovery and markedly improved limb salvage in diabetic hindlimb ischemia model relative to administration of free SP without nanodisc modification. Therefore, HDL-SP nanodisc can provide a novel strategy for the treatment of diabetic ischemia and HDL nanodisc modification could be potentially useful for the extension of plasma circulation of other labile peptides.

Synthetic High-Density Lipoprotein-Mediated Targeted Delivery of Liver X Receptors Agonist Promotes Atherosclerosis Regression.

Targeting at enhancing reverse cholesterol transport (RCT) is apromising strategy for treating atherosclerosis via infusion of reconstitute high density lipoprotein (HDL) as cholesterol acceptors or increase of cholesterol efflux by activation of macrophage liver X receptors (LXRs). However, systemic activation of LXRs triggers excessive lipogenesis in the liver and infusion of HDL downregulates cholesterol efflux from macrophages. Here we describe an enlightened strategy using phospholipid reconstituted apoA-I peptide (22A)-derived synthetic HDL (sHDL) to deliver LXR agonists to the atheroma and examine their effect on atherosclerosis regression in vivo. A synthetic LXR agonist, T0901317 (T1317) was encapsulated in sHDL nanoparticles (sHDL-T1317). Similar to the T1317 compound, the sHDL-T1317 nanoparticles upregulated the expression of ATP-binding cassette transporters and increased cholesterol efflux in macrophages in vitro and in vivo. The sHDL nanoparticles accumulated in the atherosclerotic plaques of ApoE-deficient mice. Moreover, a 6-week low-dose LXR agonist-sHDL treatment induced atherosclerosis regression while avoiding lipid accumulation in the liver. These findings identify LXR agonist loaded sHDL nanoparticles as a promising therapeutic approach to treat atherosclerosis by targeting RCT in a multifaceted manner: sHDL itself serving as both a drug carrier and cholesterol acceptor and the LXR agonist mediating upregulation of ABC transporters in the aorta.