DNA Nanostructures: Self-Adjuvant Carriers for Highly Efficient Subunit Vaccines.

Subunit vaccines based on antigen proteins or epitopes of pathogens or tumors show advantages in immunological precision and high safety, but are often limited by their low immunogenicity. Adjuvants can boost immune responses by stimulating immune cells or promoting antigen uptake by antigen presenting cells (APCs), yet existing clinical adjuvants struggle in simultaneously achieving these dual functions. Additionally, the spatial organization of antigens might be crucial to their immunogenicity. Hence, superior adjuvants should potently stimulate the immune system, precisely arrange antigens, and effectively deliver antigens to APCs. Recently, precisely organizing and delivering antigens with the unique editability of DNA nanostructures has been proposed, presenting unique abilities in significantly improving the immunogenicity of antigens. In this minireview, we will discuss the principles behind using DNA nanostructures as self-adjuvant carriers and review the latest advancements in this field. The potential and challenges associated with self-adjuvant DNA nanostructures will also be discussed.

Effect of phospholipid liposomes on prion fragment (106-128) amyloid formation.

Misfolding and aggregation of cellular prion protein (PrPc) is a major molecular process involved in the pathogenesis of prion diseases. Here, we studied the aggregation properties of a prion fragment peptide PrP(106-128). The results show that the peptide aggregates in a concentration-dependent manner in an aqueous solution and that the aggregation is sensitive to pH and the preformed amyloid seeds. Furthermore, we show that the zwitterionic POPC liposomes moderately inhibit the aggregation of PrP(106-128), whereas POPC/cholesterol (8:2) vesicles facilitate peptide aggregation likely due to the increase of the lipid packing order and membrane rigidity in the presence of cholesterol. In addition, anionic lipid vesicles of POPG and POPG/cholesterol above a certain concentration accelerate the aggregation of the peptide remarkably. The strong electrostatic interactions between the N-terminal region of the peptide and POPG may constrain the conformational plasticity of the peptide, preventing insertion of the peptide into the inner side of the membrane and thus promoting fibrillation on the membrane surface. The results suggest that the charge properties of the membrane, the composition of the liposomes, and the rigidity of lipid packing are critical in determining peptide adsorption on the membrane surface and the efficiency of the membrane in catalyzing peptide oligomeric nucleation and amyloid formation. The peptide could be used as an improved model molecule to investigate the mechanistic role of the crucial regions of PrP in aggregation in a membrane-rich environment and to screen effective inhibitors to block key interactions between these regions and membranes for preventing PrP aggregation.

Precise Epitope Organization with Self-adjuvant Framework Nucleic Acid for Efficient COVID-19 Peptide Vaccine Construction.

Peptide vaccines have advantages in easy fabrication and high safety, but their effectiveness is hampered by the poor immunogenicity of the epitopes themselves. Herein, we constructed a series of framework nucleic acids (FNAs) with regulated rigidity and size to precisely organize epitopes in order to reveal the influence of epitope spacing and carrier rigidity on the efficiency of peptide vaccines. We found that assembling epitopes on rigid tetrahedral FNAs (tFNAs) with the appropriate size could efficiently enhance their immunogenicity. Further, by integrating epitopes from SARS-CoV-2 on preferred tFNAs, we constructed a COVID-19 peptide vaccine which could induce high titers of IgG against the receptor binding domain (RBD) of SARS-CoV-2 spike protein and increase the ratio of memory B and T cells in mice. Considering the good biocompatibility of tFNAs, our research provides a new idea for developing efficient peptide vaccines against viruses and possibly other diseases.

Applications of Functional DNA Materials in Immunomodulatory Therapy.

In recent years, nanoscale or microscale functional materials derived from DNA have shown great potential for immunotherapy as superior delivery carriers. DNA nanostructures with excellent programmability and addressability enable the precise assembly of molecules or nanoparticles. DNA hydrogels have predictable structures and adjustable mechanical strength, thus being advantageous in controllable release of cargos. In addition, utilizing systematic evolution of ligands by exponential enrichment technology, a variety of DNA aptamers have been screened for specific recognition of ions, molecules, and even cells. Moreover, a wide variety of chemical modifications can further enrich the function of DNA. The unique advantages of functional DNA materials make them extremely attractive in immunomodulation. Recently, functional DNA materials-based immunotherapy has shown great potential in fighting against many diseases like cancer, viral infection, and inflammation. Therefore, in this review, we focus on discussing the progress of the applications of functional DNA materials in immunotherapy; before that, we also summarize the characteristics of the functional DNA materials descried above. Finally, we discuss the challenges and future opportunities of functional DNA materials in immunomodulatory therapy.

Framework Nucleic Acid Immune Adjuvant for Transdermal Delivery Based Chemo-immunotherapy for Malignant Melanoma Treatment.

Despite the tremendous progresses of cancer immunotherapy, its current clinical responses rate in melanoma remains to be improved. Here, we have reported a skin penetrating tetrahedral framework nucleic acid immune adjuvant (FNAIA) to transdermally deliver chemotherapy drugs into melanoma to induce the immunogenic death of tumor cells and expose tumor antigens, which with assistance of CpG oligodeoxynucleotide incorporated in FNAIA could trigger systemic tumor-specific immune responses. Compared with free CpG, FNAIA could penetrate deeper into subcutaneous tumor tissues and more effectively stimulate dendritic cell maturation. Notably, doxorubicin-loaded FNAIA locally applied on the intact skin above the melanoma could effectively inhibit the growth of mouse B16F10 melanoma and increase tumor CD8+ T cell infiltration. Moreover, combined with immune checkpoint inhibitor, the growth of distant tumors could also be effectively inhibited, suggesting that this strategy could induce systemic immune responses. Therefore, this work provides a new idea for non-invasive treatment of skin cancer.

DNA Engineered Lymphocyte-Based Homologous Targeting Artificial Antigen-Presenting Cells for Personalized Cancer Immunotherapy.

Artificial antigen-presenting cells (aAPCs) constructed by integrating T cell activation ligands on biocompatible materials hold great potential in tumor immunotherapy. However, it remains challenging to develop aAPCs, which could mimic the characteristics of natural APCs, thereby realizing antigen-specific T cells activation in vivo. Here, we report the first effort to construct natural lymphocyte-based homologous targeting aAPCs (LC-aAPCs) with lipid-DNA-mediated noninvasive live cell surface engineering. Through a predesigned bottom-up self-assembly path, we achieved natural-APC-mimicking distribution of T cell activation ligands on LC-aAPCs, which would enable the optimized T cell activation. Moreover, the lipid-DNA-mediated self-assembly occurring on lipid bilayers would not affect the functions of homing receptors expressed on lymphocyte. Therefore, such LC-aAPCs could actively migrate to peripheral lymphatic organs and then effectively activate antigen-specific T cells. Combined with an immune checkpoint inhibitor, such LC-aAPCs could effectively inhibit the growth of different tumor models. Thus, our work provides a new design of aAPCs for in vivo applications in tumor immunotherapy, and the lipid-DNA-mediated noninvasive live cell surface engineering would be a powerful tool for designing cell-based therapeutics.

Immunogenic nanomedicine based on GSH-responsive nanoscale covalent organic polymers for chemo-sonodynamic therapy.

Sonodynamic therapy (SDT) is emerging as a non-invasive strategy to eradicate tumors, but its therapeutic efficacy is still not ideal. To achieve more effective SDT, water insoluble sonosensitizer meso-5, 10, 15, 20-tetra(4-hydroxylphenyl)porphyrin (THPP) is here esterified with succinic acid conjugated oxaliplatin prodrug (Oxa(IV)SA2) and carboxyl group terminated PEG (PEG5k-COOH). The obtained covalent organic polymer (COP) of THPP-Oxa(IV)-PEG with good physiological stability, sonosensitization efficacy and glutathione (GSH) responsive oxalipatin responsive behaviors can induce effective immunogenic cancer cell death upon the ultrasound exposure. In addition, THPP-Oxa(IV)-PEG is shown to be a versatile carrier for both hydrophobic near infrared dye and radioisotope 99mTc, thereby enabling real-time tracking of its pharmacokinetics behavior under corresponding imaging facilities. Furthermore, treatment with THPP-Oxa(IV)-PEG injection and ultrasound exposure is shown to be most effectively in suppressing tumor growth, with 3 of 6 CT26 tumor bearing mice fully cured, ascribing to its high potency in eliciting profound antitumor immune responses. This work highlights a promising strategy in constructing multifunctional nanosonosensitizer as a potent immunogenic nanomedicine to enhance the treatment outcome of SDT.

ATP-Responsive Smart Hydrogel Releasing Immune Adjuvant Synchronized with Repeated Chemotherapy or Radiotherapy to Boost Antitumor Immunity.

Certain chemotherapeutics and forms of ionizing radiation can induce immunogenic cell death (ICD). If there simultaneously exist immune adjuvants within the tumor, such antitumor immunity would be further amplified. However, as clinical chemo/radiotherapies are usually repeatedly given at low individual doses, it would be impractical to administrate immune adjuvants into tumors at each dose of chemo/radiotherapies. Thus, a smart hydrogel is developed that releases immune adjuvants in response to repeatedly applied chemo-/radiotherapies. Herein, alginate is conjugated with an adenosine triphosphate (ATP)-specific aptamer, which is hybridized with immunoadjuvant CpG oligonucleotide. Upon intratumoral injection, alginate-based hydrogel is formed in situ. Interestingly, low doses of oxaliplatin or X-rays, while inducing ICD of tumor cells, could trigger release of ATP, which competitively binds with ATP-specific aptamer to trigger CpG release. Therefore, the smart hydrogel could release the immune adjuvant synchronized with low-dose repeated chemo/radiotherapies, achieving remarkable synergistic responses in eliminating established tumors, as well as immune memory to reject re-challenged tumors. Moreover, repeated radiotherapies assisted by the smart hydrogel could inhibit distant tumor metastases, especially in combination with immune checkpoint blockade. The study presents a conceptually new strategy to boost cancer immunotherapy coherent with repeated low-dose chemo-/radiotherapies following a clinically relevant manner.

Oxaliplatin-/NLG919 prodrugs-constructed liposomes for effective chemo-immunotherapy of colorectal cancer.

High expression of indoleamine 2,3-dioxygenase 1 (IDO1) is a major cause of tumor induced immunosuppression, and appears to be associated with poor prognosis in human colorectal cancer and some others. In this study, we construct a bifunctional liposome by self-assembly of oxaliplatin-prodrug (Oxa(IV)) conjugated phospholipid and alkylated NLG919 (aNLG), an IDO1 inhibitor, together with other commercial lipids. The obtained aNLG/Oxa(IV)-Lip can not only release cytotoxic oxaliplatin inside the reductive cytosol to trigger immunogenic cell death (ICD) of cancer cells, but also efficiently retard the degradation of tryptophan to immunosuppressive kynurenine via the NLG919 mediated inhibition of IDO1. Moreover, in vivo pharmacokinetic studies indicate that such aNLG/Oxa(IV)-Lip has a long blood circulation time, thereby enables highly-efficient passive tumor homing. Upon tumor accumulation, such aNLG/Oxa(IV)-Lip presents superior synergistic antitumor efficacies to both subcutaneous and orthotopic CT26 tumors, ascribing to significantly primed anti-tumor immunity of enhanced intratumoral infiltration of CD8+ T cells, scretion of cytotoxic cytokines and downregulation of immunosuppressive regulatory T cells. This work highlights that such bifunctional aNLG/Oxa(IV)-Lip is a potent candidate for future clinical translation owing to its excellent biocompatibility and high therapeutic efficacy.

DNA-Edited Ligand Positioning on Red Blood Cells to Enable Optimized T Cell Activation for Adoptive Immunotherapy.

Artificial antigen presenting cells (aAPCs) with surface-anchored T cell activating ligands hold great potential in adoptive immunotherapy. However, it remains challenging to precisely control the ligand positioning on those platforms using conventional bioconjugation chemistry. Utilizing DNA-assisted bottom-up self-assembly, we were able to precisely control both lateral and vertical distributions of T cell activation ligands on red blood cells (RBCs). The clustered lateral positioning of the peptide-major histocompatibility complex (pMHC) on RBCs with a short vertical distance to the cell membrane is favorable for more effective T cell activation, likely owing to their better mimicry of natural APCs. Such optimized RBC-based artificial APCs can stimulate T cell proliferation in vivo and effectively inhibit tumor growth with adoptive immunotherapy. DNA technology is thus a unique tool to precisely engineer the cell membrane interface and tune cell-cell interactions, which is promising for applications such as immunotherapy.

Cell-Penetrating Peptide Enhanced Antigen Presentation for Cancer Immunotherapy.

The development of effective cancer vaccines is an important direction in the area of cancer immunotherapy. Although certain types of preventive cancer vaccines have already been used in the clinic, therapeutic cancer vaccines for treatment of already established tumors are still in high demand. In this study, we develop a new type of cancer vaccine by mixing cell-penetrating peptide (CPP) conjugated antigen as the enhanced antigen, together with CpG as the immune adjuvant. A special CPP, cytosol-localizing internalization peptide 6 (CLIP6), which has the ability to enter cells exclusively via a nonendosomal mechanism, i.e., direct translocation across the cell membrane, is conjugated with model antigen ovalbumin (OVA). Compared to naked OVA, the obtained CLIP6-OVA conjugates show greatly increased uptake by dendritic cells (DCs) and, more importantly, remarkably enhanced antigen cross-presentation, eliciting stronger cytotoxic T lymphocyte (CTL) mediated immune responses with the help of CpG. This CLIP6-OVA/CpG formulation offers effective protection for mice against challenged B16-OVA tumors, and is able to further function as a therapeutic vaccine, which, in combination with immune checkpoint blockade therapy, can significantly suppress the already-established tumors. Such a CLIP6-based cancer vaccine developing strategy shows promising potential toward clinical practice owing to its features of easy preparation, low cost, and remarkable biocompatibility.