Vaccines for Non-Viral Cancer Prevention

Cancer vaccines are a type of immune therapy that seeks to modulate the host’s immune system to induce durable and protective immune responses against cancer-related antigens. The little clinical success of therapeutic cancer vaccines is generally attributed to the immunosuppressive tumor microenvironment at late-stage diseases. The administration of cancer-preventive vaccination at early stages, such as pre-malignant lesions or even in healthy individuals at high cancer risk could increase clinical efficacy by potentiating immune surveillance and pre-existing specific immune responses, thus eliminating de novo appearing lesions or maintaining equilibrium. Indeed, research focus has begun to shift to these approaches and some of them are yielding encouraging outcomes.     

Protein-Based Nanoparticle Vaccines for SARS-CoV-2

The pandemic caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has upended healthcare systems and economies around the world. Rapid understanding of the structural biology and pathogenesis of SARS-CoV-2 has allowed the development of emergency use or FDA-approved vaccines and various candidate vaccines. Among the recently developed SARS-CoV-2 candidate vaccines, natural protein-based nanoparticles well suited for multivalent antigen presentation and enhanced immune stimulation to elicit potent humoral and cellular immune responses are currently being investigated. This mini-review presents recent innovations in protein-based nanoparticle vaccines against SARS-CoV-2. The design and strategy of displaying antigenic domains, including spike protein, receptor-binding domain (RBD), and other domains on the surface of various protein-based nanoparticles and the performance of the developed nanoparticle-based vaccines are highlighted. In the final part of this review, we summarize and discuss recent advances in clinical trials and provide an outlook on protein-based nanoparticle vaccines.     

TLR2 Deficiency Exacerbates Imiquimod-Induced Psoriasis-Like Skin Inflammation through Decrease in Regulatory T Cells and Impaired IL-10 Production

Emerging evidence has demonstrated that Toll-like receptors (TLRs) are associated with autoimmune diseases. In this study, we investigated the role of TLR2 in psoriasis using imiquimod-induced psoriasis-like dermatitis. Although TLR2 signaling is known to play a critical role in the induction of proinflammatory cytokines by immune cells, such as dendritic cells (DCs), macrophages, and monocytes, TLR2 deficiency unexpectedly exacerbated psoriasiform skin inflammation. Importantly, messenger RNA (mRNA) levels of Foxp-3 and IL-10 in the lesional skin were significantly decreased in TLR2 KO mice compared with wild-type mice. Furthermore, flow cytometric analysis of the lymph nodes revealed that the frequency of regulatory T cells (Tregs) among CD4-positive cells was decreased. Notably, stimulation with Pam3CSK4 (TLR2/1 ligand) or Pam2CSK4 (TLR2/6 ligand) increased IL-10 production from Tregs and DCs and the proliferation of Tregs. Finally, adoptive transfer of Tregs from wild-type mice reduced imiquimod-induced skin inflammation in TLR2 KO mice. Taken together, our results suggest that TLR2 signaling directly enhances Treg proliferation and IL-10 production by Tregs and DCs, suppressing imiquimod-induced psoriasis-like skin inflammation. Enhancement of TLR2 signaling may be a new therapeutic strategy for psoriasis.     

Multivalent,Stabilized Mannose-6-Phosphates for the Targeted Delivery of Toll-Like Receptor Ligands and Peptide Antigens

Mannose-6-phosphate (M6P) is recognized by the mannose-6-phosphate receptor and plays an important role in the transport of cargo to the endosomes, making it an attractive tool to improve endosomal trafficking of vaccines. We describe herein the assembly of peptide antigen conjugates carrying clusters of mannose-6-C-phosphonates (M6Po). The M6Po's are stable M6P mimics that are resistant to cleavage of the phosphate group by endogenous phosphatases. Two different strategies for the incorporation of the M6Po clusters in the conjugate have been developed: the first relies on a “post-assembly” click approach employing an M6Po bearing an alkyne functionality; the second hinges on an M6Po C-glycoside amino acid building block that can be used in solid-phase peptide synthesis. The generated conjugates were further equipped with a TLR7 ligand to stimulate dendritic cell (DC) maturation. While antigen presentation is hindered by the presence of the M6Po clusters, the incorporation of the M6Po clusters leads to increased activation of DCs, thus demonstrating their potential in improving vaccine adjuvanticity by intraendosomally active TLR ligands.     

Targeting Death Receptor TRAIL-R2 by Chalcones for TRAIL-Induced Apoptosis in Cancer Cells

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in cancer cells without toxicity to normal cells. TRAIL binds to death receptors, TRAIL-R1 (DR4) and TRAIL-R2 (DR5) expressed on cancer cell surface and activates apoptotic pathways. Endogenous TRAIL plays an important role in immune surveillance and defense against cancer cells. However, as more tumor cells are reported to be resistant to TRAIL mediated death, it is important to search for and develop new strategies to overcome this resistance. Chalcones can sensitize cancer cells to TRAIL-induced apoptosis. We examined the cytotoxic and apoptotic effects of TRAIL in combination with four chalcones: chalcone, isobavachalcone, licochalcone A and xanthohumol on HeLa cancer cells. The cytotoxicity was measured by MTT and LDH assays. The apoptosis was detected using annexin V-FITC staining by flow cytometry and fluorescence microscopy. Death receptor expression was analyzed using flow cytometry. The decreased expression of death receptors in cancer cells may be the cause of TRAIL-resistance. Chalcones enhance TRAIL-induced apoptosis in HeLa cells through increased expression of TRAIL-R2. Our study has indicated that chalcones augment the antitumor activity of TRAIL and confirm their cancer chemopreventive properties.     

Beta 2-Adrenergic Receptor in Circulating Cancer-Associated Cells Predicts for Increases in Stromal Macrophages in Circulation and Patient Survival in Metastatic Breast Cancer

The usage of beta blockers in breast cancer (BC) patients is implicated in the reduction in distant metastases, cancer recurrence, and cancer mortality. Studies suggest that the adrenergic pathway is directly involved in sympathetic-driven hematopoietic activation of pro-tumor microenvironmental proliferation and tumor cell trafficking into the circulation. Cancer-associated macrophage-like cells (CAMLs) are pro-tumor polynucleated monocytic cells of hematopoietic origin emanating from tumors which may aid in circulating tumor cell (CTC) dissemination into the circulation. We examined the linkage between Beta-2 adrenergic receptor (B2AR) signaling in CAMLs and CTCs by establishing expression profiles in a model BC cell line (MDA-MB-231). We compared the model to CAMLs and CTCs found in patents. Although internalization events were observed in patients, differences were found in the expression of B2AR between the tumor cell lines and the CAMLs found in patients. High B2AR expression on patients’ CAMLs was correlated with significantly more CAMLs in the circulation (p = 0.0093), but CTCs had no numerical relationship (p = 0.1565). High B2AR CAML expression was also significantly associated with a larger size of CAMLs (p = 0.0073), as well as being significantly associated with shorter progression-free survival (p = 0.0097) and overall survival (p = 0.0265). These data suggest that B2AR expression on CAMLs is closely related to the activation, intravasation, and growth of CAMLs in the circulation.     

Toll-like Receptor 2 Is Associated with the Immune Response,Apoptosis, and Angiogenesis in the Mammary Glands of Dairy Cows with Clinical Mastitis

Toll-like receptor 2 (TLR2) plays a crucial role in bacterial recognition and the host immune response during infection. However, its function and downstream biological processes (BPs) in the mammary glands (MGs) of Holstein cows with clinical mastitis (CM) are not fully understood. This study aimed to comprehensively identify the BPs and differentially expressed proteins (DEPs) associated with the bacterial response and TLR2 using data-independent acquisition (DIA) proteomic data. A possible mechanism for the action of TLR2 was proposed, and the results suggested that the expression levels of TLR2 and caspase 8 (CASP8) were positively correlated with the apoptosis of MGs. The expression patterns of TLR2 and TEK receptor tyrosine kinase 2 (Tie2) were negatively correlated with angiogenesis. These results indicated that TLR2 might promote apoptosis in mammary epithelial cells (MECs) and vascular endothelial cells (VECs) via upregulation of CASP8 expression, and inhibition of angiogenesis in VECs via downregulation of Tie2 expression in dairy cows with CM. In conclusion, TLR2 is associated with inflammation, apoptosis, and angiogenesis in the MGs of dairy cows with bacteria-induced mastitis. These results contribute to a deeper understanding of the pathogenic mechanisms and provide the knowledge needed for developing the prevention and treatment of dairy mastitis.     

Helicobacter pylori Neutrophil-Activating Protein Directly Interacts with and Activates Toll-like Receptor 2 to Induce the Secretion of Interleukin-8 from Neutrophils and ATRA-Induced Differentiated HL-60 Cells

Helicobacter pylori neutrophil-activating protein (HP-NAP)-induced production of reactive oxygen species (ROS) by neutrophils and monocytes is regulated by pertussis toxin (PTX)-sensitive G proteins, whereas HP-NAP-induced cytokine secretion by monocytes is mediated by Toll-like receptor 2 (TLR2). However, it is unclear whether TLR2 participates in HP-NAP-induced cytokine secretion by neutrophils. Here, all-trans retinoic acid (ATRA)-induced differentiated HL-60 cells were first employed as a neutrophil model to investigate the molecular mechanisms underlying neutrophil responses to HP-NAP. HP-NAP-induced ROS production in ATRA-induced differentiated HL-60 cells is mediated by the PTX-sensitive heterotrimeric G protein-dependent activation of extracellular signal-regulated kinase 1/2 and p38-mitogen-activated protein kinase, which is consistent with the findings reported for human neutrophils. Next, whether TLR2 participated in HP-NAP-induced secretion of interleukin-8 (IL-8) was investigated in neutrophils and ATRA-induced differentiated HL-60 cells. In both cells, TLR2 participated in HP-NAP-induced IL-8 secretion but not HP-NAP-induced ROS production. Interestingly, PTX-sensitive G proteins also contributed to the HP-NAP-induced secretion of IL-8 from neutrophils and the differentiated HL-60 cells. Our ELISA-based binding assay further revealed the competitive binding of Pam₃CSK₄, a TLR2 agonist, and HP-NAP to TLR2, which suggests the presence of specific and direct interactions between HP-NAP and TLR2. Thus, HP-NAP directly interacts with and activates TLR2 to induce IL-8 secretion in neutrophils and ATRA-induced differentiated HL-60 cells.     

Cancer Vaccines,Treatment of the Future: With Emphasis on HER2-Positive Breast Cancer

Breast cancer is one of the leading causes of death in women. With improvements in early-stage diagnosis and targeted therapies, there has been an improvement in the overall survival rate in breast cancer over the past decade. Despite the development of targeted therapies, tyrosine kinase inhibitors, as well as monoclonal antibodies and their toxin conjugates, all metastatic tumors develop resistance, and nearly one-third of HER2+ breast cancer patients develop resistance to all these therapies. Although antibody therapy has shown promising results in breast cancer patients, passive immunotherapy approaches have limitations and need continuous administration over a long period. Vaccine therapy introduces antigens that act on cancer cells causing prolonged activation of the immune system. In particular, cancer relapse could be avoided due to the presence of a longer period of immunological memory with an effective vaccine that can protect against various tumor antigens. Cancer vaccines are broadly classified as preventive and therapeutic. Preventive vaccines are used to ward off any future infections and therapeutic vaccines are used to treat a person with active disease. In this article, we provided details about the tumor environment, different types of vaccines, their advantages and disadvantages, and the current status of various vaccine candidates with a focus on vaccines for breast cancer. Current data indicate that therapeutic vaccines themselves have limitations in terms of efficacy and are used in combination with other chemotherapeutic or targeting agents. The majority of breast cancer vaccines are undergoing clinical trials and the next decade will see the fruitfulness of breast cancer vaccine therapy.     

Dendritic Cells Pulsed with Cytokine-Adjuvanted Tumor Membrane Vesicles Inhibit Tumor Growth in HER2-Positive and Triple Negative Breast Cancer Models

Dendritic cells (DCs) are the most effective antigen presenting cells for the development of T cell responses. The only FDA approved DC-based immunotherapy to date is Sipuleucel-T, which utilizes a fusion protein to stimulate DCs ex vivo with GM-CSF and simultaneously deliver the antigen PAP for prostate cancer. This approach is restricted by the breadth of immunity elicited to a single antigen, and to cancers that have a defined tumor associated antigen. Other multi-antigen approaches have been restricted by poor efficacy of vaccine adjuvants. We have developed a vaccine platform that consists of autologous DCs pulsed with cytokine-adjuvanted tumor membrane vesicles (TMVs) made from tumor tissue, that encapsulate the antigenic landscape of individual tumors. Here we test the efficacy of DCs pulsed with TMVs incorporated with glycolipid-anchored immunostimulatory molecules (GPI-ISMs) in HER2-positive and triple negative breast cancer murine models. Pulsing of DCs with TMVs containing GPI-ISMs results in superior uptake of vesicles, DC activation and cytokine production. Adaptive transfer of TMV-pulsed DCs to tumor bearing mice results in the inhibition of tumor growth, reduction in lung metastasis, and an increase in immune cell infiltration into the tumors. These observations suggest that DCs pulsed with TMVs containing GPI-GM-CSF and GPI-IL-12 can be further developed to be used as a personalized immunotherapy platform for cancer treatment.     

Synthetic Indolactam V Analogues as Inhibitors of PAR2‐Induced Calcium Mobilization in Triple‐Negative Breast Cancer Cells

Human proteinase‐activated receptor 2 (PAR2), a transmembrane G‐protein‐coupled receptor (GPCR), is an attractive target for a novel anticancer therapy, as it plays a critical role in cell migration and invasion. Selective PAR2 inhibitors therefore have potential as anti‐metastatic drugs. Knowing that the natural product teleocidin A2 is able to inhibit PAR2 in tumor cells, the goal of the present study was to elaborate structure–activity relationships and to identify potent PAR2 inhibitors with lower activity against the adverse target, protein kinase C (PKC). For this purpose, an efficient gram‐scale total synthesis of indolactam V (i.e., the parent structure of all teleocidins) was developed, and a library of derivatives was prepared. Some compounds were indeed found to exhibit high potency as PAR2 inhibitors at low nanomolar concentrations with improved selectivity (relative to teleocidin A2). The pseudopeptidic fragment bridging the C3 and C4 positions of the indole core proved to be essential for target binding, whereas activity and target selectivity depends on the substituents at N1 or C7. This study revealed novel derivatives that show high efficacy in PAR2 antagonism combined with increased selectivity.     

Tryptophan Metabolites at the Crossroad of Immune-Cell Interaction via the Aryl Hydrocarbon Receptor: Implications for Tumor Immunotherapy

The Aryl hydrocarbon receptor (AhR) is a critical regulator of both innate and adaptive immune responses, with potent immunomodulatory effects that makes this receptor an attractive molecular target for novel therapeutics. Accumulating evidence indicates that diverse—both host’s and microbial—tryptophan metabolites profoundly regulate the immune system in the host via AhR, promoting either tolerance or immunity, largely as a function of the qualitative and quantitative nature of the metabolites being contributed by either source. Additional findings indicate that host and microbiota-derived tryptophan metabolic pathways can influence the outcome of immune responses to tumors. Here, we review recent studies on the role and modalities of AhR activation by various ligands, derived from either host-cell or microbial-cell tryptophan metabolic pathways, in the regulation of immune responses. Moreover, we highlight potential implications of those ligands and pathways in tumor immunotherapy, with particular relevance to checkpoint-blockade immune intervention strategies.     

Strategies for Synthesizing and Enhancing the Immune Response of Cancer Vaccines Based on MUC1 Glycopeptide Antigens

Cancer vaccines are based on a vaccinology strategy whereby the patient's immune system is harnessed to induce a specific immune response to kill cancer cells and comprises two categories: prophylactic and therapeutic. Glycoprotein mucin 1 (MUC1), which is overexpressed and poorly glycosylated on cancer cells, is one of the most promising candidates for the development of new cancer vaccines. However, it should be noted that mucin-like glycopeptides are poorly immunogenic and unable to elicit effective and long-lasting immune responses. Therefore, MUC1-derived tumor antigens need to be conjugated with immune activators. This review focuses on the synthesis of MUC1 glycopeptides, provides an overview of recently advanced designs of vaccines based on MUC1, and compares the advantages and disadvantages of the various strategies devised to date.     

4-甲基-2-肼基苯并噻唑合成工艺研究

研究了一种4-甲基-2-肼基苯并噻唑(简称:肼基苯并噻唑)的合成新工艺,该工艺以水代替乙二醇作为反应介质,以适当比例无机盐、肼盐、水合肼组成的复合体系与4-甲基-2-氨基苯并噻唑(简称:氨基苯并噻唑)反应,用有机萃取剂分离产物和母液,母液循环套用,有机萃取剂直接单釜化操作合成三环唑.该工艺操作简单成本低,收率含量指标与溶剂法相当,宜于工业化推广.     

4,6-二甲基-2-巯基嘧啶合成方法的改进

改进了4,6-二甲基-2-巯基嘧啶的合成方法,探讨了反应温度、反应时间和溶剂量对产率的影响.通过正交实验确定优化合成条件为:反应温度65℃、反应时间5 h、溶剂甲醇量35 mL(乙酰丙酮∶硫脲=0.08 mol∶0.08 mol).改进后的合成方法具有耗时短及操作简便的特点,产率可达85%.     

Regulatory T Cells with Additional COX-2 Expression Are Independent Negative Prognosticators for Vulvar Cancer Patients

Vulvar cancer incidence numbers have been steadily rising over the past decades. In particular, the number of young patients with vulvar cancer has recently increased. Therefore, the need to identify new prognostic factors and, in addition, therapeutic options for vulvar carcinoma is more apparent. The aim of this study was to analyze the influx of COX-2 positive tumor-infiltrating lymphocytes and monocytes and their influence on prognosis. Using subtyping by immunofluorescence, the majority of COX-2 expressing immune cells were identified as FOXP3-positive regulatory T cells. In addition, peri- and intra-tumoral macrophages in the same tumor tissue were detected simultaneously as M2-polarized macrophages. COX-2 positive immune cells were independent negative prognostic markers in long-term overall survival of patients with vulvar cancer. These results show an influence of immune cell infiltration for vulvar carcinoma patients. Immune cell infiltration and immune checkpoint expression may, therefore, become interesting targets for further research on new vulvar cancer treatment strategies.     

Novel and Selective Fluorescent σ2‐Receptor Ligand with a 3,4‐Dihydroisoquinolin‐1‐one Scaffold: A Tool to Study σ2 Receptors in Living Cells

Although sigma‐2 (σ₂) receptors are still enigmatic proteins, they are promising targets for tumor treatment and diagnosis. With the aim of clarifying their role in oncology, we developed a σ₂‐selective fluorescent tracer (compound 5 ) as a specific tool to study σ₂ receptors. By using flow cytometry with 5 , we performed competition binding studies on three different cell lines where we also detected the content of the σ₂ receptors, avoiding the inconvenient use of radioligands. Comparison with a previously developed mixed σ₁/σ₂ fluorescent tracer ( 1 ) also allowed for the detection of σ₁ receptors within these cells. Results obtained by flow cytometry with tracers 1 and 5 were confirmed by standard methods (western blot for σ₁, and Scatchard analysis for σ₂ receptors). Thus, we have produced powerful new tools for research on the σ whose reliability and adaptability to a number of fluorescence techniques will be useful to elucidate the roles of σ receptors in oncology.     

Thapsigargin-Stimulated LAD2 Human Mast Cell Line Is a Potent Cellular Adjuvant for the Maturation of Monocyte-Derived Dendritic Cells for Adoptive Cellular Immunotherapy

The preparation of dendritic cells (DCs) for adoptive cellular immunotherapy (ACI) requires the maturation of ex vivo-produced immature(i) DCs. This maturation ensures that the antigen presentation triggers an immune response towards the antigen-expressing cells. Although there is a large number of maturation agents capable of inducing strong DC maturation, there is still only a very limited number of these agents approved for use in the production of DCs for ACI. In seeking novel DC maturation agents, we used differentially activated human mast cell (MC) line LAD2 as a cellular adjuvant to elicit or modulate the maturation of ex vivo-produced monocyte-derived iDCs. We found that co-culture of iDCs with differentially activated LAD2 MCs in serum-containing media significantly modulated polyinosinic:polycytidylic acid (poly I:C)-elicited DC maturation as determined through the surface expression of the maturation markers CD80, CD83, CD86, and human leukocyte antigen(HLA)-DR. Once iDCs were generated in serum-free conditions, they became refractory to the maturation with poly I:C, and the LAD2 MC modulatory potential was minimized. However, the maturation-refractory phenotype of the serum-free generated iDCs was largely overcome by co-culture with thapsigargin-stimulated LAD2 MCs. Our data suggest that differentially stimulated mast cells could be novel and highly potent cellular adjuvants for the maturation of DCs for ACI.