Depletion of regulatory T cells enhancing the anti-tumor effect of in situ vaccination in solid tumors.

The emergence of immune checkpoint inhibitors (ICIs) has revolutionized the clinical treatment for tumor. However, the low response rate of ICIs remains the major obstacle for curing patients and effective approaches for patients with primary or secondary resistance to ICIs remain lacking. In this study, immune stimulating agent unmethylated CG-enriched (CpG) oligodeoxynucleotide (ODN) was locally injected into the tumor to trigger a robust immune response to eradicate cancer cells, while anti-CD25 antibody was applied to remove immunosuppressive regulatory T cells, which further enhanced the host immune activity to attack tumor systematically. The combination of CpG and anti-CD25 antibody obtained notable regression in mouse melanoma model. Furthermore, rechallenge of tumor cells in the xenograft model has resulted in smaller tumor volume, which demonstrated that the combinational treatment enhanced the activity of memory T cells. Remarkably, this combinational therapy presented significant efficacy on multiple types of tumors as well and was able to prevent relapse of tumor partially. Taken together, our combinational immunotherapy provides a new avenue to enhance the clinical outcomes of patients who are insensitive or resistant to ICIs treatments.

Small EV-based delivery of CpG ODNs for melanoma postsurgical immunotherapy.

Blocking programmed cell death protein 1 (PD-1) is an effective therapeutic strategy for melanoma. However, patients often develop tumor recurrence postoperatively due to the low response rate to the anti-PD-1 antibody (aPD-1). In this study, we developed an in situ sprayable fibrin gel that contains cytosine-guanine oligodeoxynucleotides (CpG ODNs)-modified ovalbumin (OVA) antigen-expressing bone marrow dendritic cell (DC)-derived small extracellular vesicles (DC-sEVs) and aPD-1. CpG ODNs can activate DCs, which have potent immunostimulatory effects, by stimulating both the maturation and activation of tumor-infiltrating dendritic cells (TIDCs) and DCs in tumor-draining lymph nodes (TDLNs). In addition, DC-sEVs can deliver OVA to the same DCs, leading to the specific expression of tumor antigens by antigen-presenting cells (APCs). In brief, the unique synergistic combination of aPD-1 and colocalized delivery of immune adjuvants and tumor antigens enhances antitumor T-cell immunity, not only in the tumor microenvironment (TME) but also in TDLNs. This effectively attenuates local tumor recurrence and metastasis. Our results suggest that dual activation by CpG ODNs prolongs the survival of mice and decreases the recurrence rate in an incomplete tumor resection model, providing a promising approach to prevent B16-F10-OVA melanoma tumor recurrence and metastasis.

The potential role of CpG oligodeoxynucleotides on diabetic cardiac autonomic neuropathy mediated by P2Y12 receptor in rat stellate ganglia.

Cardiac autonomic neuropathy has a high prevalence in type 2 diabetes, which increases the risk of cardiovascular system disorders. CpG oligodeoxynucleotide (CpG-ODN), a Toll-like receptor 9 (TLR9) ligand, has been shown to have cardioprotection and cellular protection. Our previous work showed that P2Y12 in stellate ganglia (SG) is involved in the process of diabetic cardiac autonomic neuropathy (DCAN). Here, we aim to investigate whether CpG-ODN 1826 plays a protective role in DCAN and whether this beneficial protection involves regulation of the P2Y12-mediated cardiac sympathetic injury. Our results revealed that CpG-ODN 1826 activated TLR9 receptor, improved the abnormal blood pressure (BP), heart rate (HR), heart rate variability (HRV) and sympathetic nerve discharge (SND) activity in diabetic rats and reduced the up-regulated NF-κB, P2Y12 receptor, TNF-α and IL-1ß in SG. Meanwhile, CpG-ODN 1826 significantly decreased the elevated ATP, nuclear receptor coactivator 4 (NCOA4), iron, ROS and MDA levels and increased GPX4 and GSH levels. In addition, CpG-ODN 1826 contributes to maintain normalization of mitochondrial structure in SG. Overall, CpG-ODN 1826 alleviates the sympathetic excitation and abnormal neuron-glial signal communication via activating TLR9 receptors to achieve a balance of autonomic activity and relieve the DCAN in rats. The mechanism may involve the regulation of P2Y12 receptor in SG by reducing ATP release and NF-κB expression, which counteract neuroinflammation and ferroptosis mediated by activated P2Y12 in SG.

STAT3/NF­κB decoy oligodeoxynucleotides inhibit atherosclerosis through regulation of the STAT/NF­κB signaling pathway in a mouse model of atherosclerosis.

Atherosclerosis is a progressive chronic inflammatory condition that is the cause of most cardiovascular and cerebrovascular diseases. The transcription factor nuclear factor­κB (NF­κB) regulates a number of genes involved in the inflammatory responses of cells that are critical to atherogenesis, and signal transducer and activator of transcription (STAT)3 is a key transcription factor in immunity and inflammation. Decoy oligodeoxynucleotides (ODNs) bind to sequence­specific transcription factors and limit gene expression by interfering with transcription in vitro and in vivo. The present study aimed to investigate the beneficial functions of STAT3/NF­κB decoy ODNs in liposaccharide (LPS)­induced atherosclerosis in mice. Atherosclerotic injuries of mice were induced via intraperitoneal injection of LPS and the mice were fed an atherogenic diet. Ring­type STAT3/NF­κB decoy ODNs were designed and administered via an injection into the tail vein of the mice. To investigate the effect of STAT3/NF­κB decoy ODNs, electrophoretic mobility shift assay, western blot analysis, histological analysis with hematoxylin and eosin staining, Verhoeff­Van Gieson and Masson's trichrome staining were performed. The results revealed that STAT3/NF­κB decoy ODNs were able to suppress the development of atherosclerosis by attenuating morphological changes and inflammation in atherosclerotic mice aortae, and by reducing pro­inflammatory cytokine secretion through inhibition of the STAT3/NF­κB pathway. In conclusion, the present study provided novel insights into the antiatherogenic molecular mechanism of STAT3/NF­κB decoy ODNs, which may serve as an additional therapeutic intervention to combat atherosclerosis.

BAX as the mediator of C-MYC sensitizes acute lymphoblastic leukemia to TLR9 agonists.

BACKGROUND: The prognosis of B-cell acute lymphoblastic leukemia (B-ALL) has improved significantly with current first-line therapy, although the recurrence of B-ALL is still a problem. Toll-like receptor 9 (TLR9) agonists have shown good safety and efficiency as immune adjuvants. Apart from their immune regulatory effect, the direct effect of TLR9 agonists on cancer cells with TLR9 expression cannot be ignored. However, the direct effect of TLR9 agonists on B-ALL remains unknown. METHODS: We discussed the relationship between TLR9 expression and the clinical characteristics of B-ALL and explored whether CpG 685 exerts direct apoptotic effect on B-ALL without inhibiting normal B-cell function. By using western blot, co-immunoprecipitation, immunofluorescence co-localization, and chromatin immunoprecipitation, we explored the mechanism of the apoptosis-inducing effect of CpG 685 in treating B-ALL cells. By exploring the mechanism of CpG 685 on B-ALL, the predictive biomarkers of the efficacy of CpG 685 in treating B-ALL were explored. These efficiencies were also confirmed in mouse model as well as clinical samples. RESULTS: High expression of TLR9 in B-ALL patients showed good prognosis. C-MYC-induced BAX activation was the key to the effect of CpG oligodeoxynucleotides against B-ALL. C-MYC overexpression promoted P53 stabilization, enhanced Bcl-2 associated X-protein (BAX) activation, and mediated transcription of the BAX gene. Moreover, combination therapy using CpG 685 and imatinib, a BCR-ABL kinase inhibitor, could reverse resistance to CpG 685 or imatinib alone by promoting BAX activation and overcoming BCR-ABL1-independent PI3K/AKT activation. CONCLUSION: TLR9 is not only a prognostic biomarker but also a potential target for B-ALL therapy. CpG 685 monotherapy might be applicable to Ph- B-ALL patients with C-MYC overexpression and without BAX deletion. CpG 685 may also serve as an effective combinational therapy against Ph+ B-ALL.

Modulation of the Inflammatory Response by Pre-emptive Administration of IMT504 Reduces Postoperative Pain in Rats and has Opioid-Sparing Effects.

Despite the available knowledge on underlying mechanisms and the development of several therapeutic strategies, optimal management of postoperative pain remains challenging. This preclinical study hypothesizes that, by promoting an anti-inflammatory scenario, pre-emptive administration of IMT504, a noncoding, non-CpG oligodeoxynucleotide with immune modulating properties, will reduce postincisional pain, also facilitating therapeutic opioid-sparing. Male adult Sprague-Dawley rats with unilateral hindpaw skin-muscle incision received pre-emptive (48 and 24 hours prior to surgery) or postoperative (6 hours after surgery) subcutaneous vehicle (saline) or IMT504. Various groups of rats were prepared for pain-like behavior analyses, including subgroups receiving morphine or naloxone, as well as for flow-cytometry or quantitative RT-PCR analyses of the spleen and hindpaws (for analysis of inflammatory phenotype). Compared to vehicle-treated rats, pre-emptive IMT504 significantly reduced mechanical allodynia by 6 hours after surgery, and accelerated recovery of basal responses from 72 hours after surgery and onwards. Cold allodynia was also reduced by IMT504. Postoperative administration of IMT504 resulted in similar positive effects on pain-like behavior. In IMT504-treated rats, 3 mg/kg morphine resulted in comparable blockade of mechanical allodynia as observed in vehicle-treated rats receiving 10 mg/kg morphine. IMT504 significantly increased hindpaw infiltration of mesenchymal stem cells, CD4+T and B cells, and caused upregulated or downregulated transcript expressions of interleukin-10 and interleukin-1ß, respectively. Also, IMT504 treatment targeted the spleen, with upregulated or downregulated transcript expressions, 6 hours after incision, of interleukin-10 and interleukin-1ß, respectively. Altogether, pre-emptive or postoperative IMT504 provides protection against postincisional pain, through participation of significant immunomodulatory actions, and exhibiting opioid-sparing effects. PERSPECTIVE: This preclinical study introduces the noncoding non-CpG oligodeoxynucleotide IMT504 as a novel modulator of postoperative pain and underlying inflammatory events. The opioid-sparing effects observed for IMT504 appear as a key feature that could contribute, in the future, to reducing opioid-related adverse events in patients undergoing surgical intervention.

Immunotherapy for lung cancer combining the oligodeoxynucleotides of TLR9 agonist and TGF-ß2 inhibitor.

Tumor immunotherapies have shown promising antitumor effects, especially immune checkpoint inhibitors (ICIs). However, only 12.46% of the patients benefit from the ICIs, the rest of them shows limited effects on ICIs or even accelerates the tumor progression due to the lack of the immune cell infiltration and activation in the tumor microenvironment (TME). In this study, we administrated a combination of Toll-like receptor 9 (TLR9) agonist CpG ODN and Transforming growth factor-ß2 (TGF-ß2) antisense oligodeoxynucleotide TIO3 to mice intraperitoneally once every other day for a total of four injections, and the first injection was 24 h after LLC cell inoculation. We found that the combination induced the formation of TME toward the enrichment and activation of CD8+ T cells and NK cells, accompanied with a marked decrease of TGF-ß2. The combined therapy also effectively inhibited the tumor growth and prolonged the survival of the mice, even protected the tumor-free mice from the tumor re-challenge. Both of CpG ODN and TIO3 are indispensable, because replacing CpG ODN with TLR9 inhibitor CCT ODN showed no antitumor effect, CpG ODN or TIO3 alone did not lead to ideal antitumor results. This effect was possibly initiated by the activation of dendritic cells at the tumor site. This systemic antitumor immunotherapy with a combination of the two oligonucleotides (an immune stimulant and an immunosuppressive cytokine inhibitor) before the tumor formation may provide a novel strategy for clinical prevention of the postoperative tumor recurrence.

DNA Nanoflowers' Amelioration of Lupus Symptoms in Mice via Blockade of TLR7/9's Signal.

Inhibitory oligodeoxynucleotides (INH-ODN) can exert an immunomodulatory effect to specifically block TLR7 and TLR9 signaling in systemic lupus erythematosus (SLE). To extend the half-life of INH-ODN in vivo, the phosphorothioate backbone, instead of the native phosphodiester, is preferred due to its strong resistance against nuclease degradation. However, its incomplete degradation in vivo may lead to potential risk. To solve these problems and enhance the blockage of TLR7 and TLR9, we prepared highly compressed DNA nanoflowers with prolonged native DNA backbones and repeated INH-ODN motifs. Three therapeutic types of nanoflower, incorporating INH-ODN sequences, including IRS 661, IRS 869, and IRS 954, were prepared by rolling circle amplification and were subcutaneously injected into MRL/lpr mice. The TLR7 blocker of the IRS 661 nanoflower and the TLR9 antagonist of the IRS 869 nanoflower could decrease autoantibodies, reduce cytokine secretion, and alleviate lupus nephritis in mice. However, the IRS 954 nanoflower, the TLR7 and TLR9 dual antagonist, did not have additive or opposing effects on lupus nephritis but only showed a decrease in serum IFNα, suggesting that the TLR7 and TLR9 antagonist may have a competition mechanism or signal-dependent switching relationship. INH-ODN nanoflowers were proposed as a novel and potential therapeutic nucleic acids for SLE.

MF59 Promoted the Combination of CpG ODN1826 and MUC1-MBP Vaccine-Induced Antitumor Activity Involved in the Enhancement of DC Maturation by Prolonging the Local Retention Time of Antigen and Down-Regulating of IL-6/STAT3.

Our previous study found that CpG oligodeoxynucleotides 1826 (CpG 1826), combined with mucin 1 (MUC1)-maltose-binding protein (MBP) (M-M), had certain antitumor activity. However, this combination is less than ideal for tumor suppression (tumors vary in size and vary widely among individuals), with a drawback being that CpG 1826 is unstable. To solve these problems, here, we evaluate MF59/CpG 1826 as a compound adjuvant with M-M vaccine on immune response, tumor suppression and survival. The results showed that MF59 could promote the CpG 1826/M-M vaccine-induced tumor growth inhibition and a Th1-prone cellular immune response, as well as reduce the individual differences of tumor growth and prolonged prophylactic and therapeutic mouse survival. Further research showed that MF59 promotes the maturation of DCs stimulated by CpG1826/M-M, resulting in Th1 polarization. The possible mechanism is speculated to be that MF59 could significantly prolong the retention time of CpG 1826, or the combination of CpG 1826 and M-M, as well as downregulate IL-6/STAT3 involved in MF59 combined CpG 1826-induced dendritic cell maturation. This study clarifies the utility of MF59/CpG 1826 as a vaccine compound adjuvant, laying the theoretical basis for the development of a novel M-M vaccine.

Intratumoral Therapy to Make a "Cold" Tumor "Hot": The Jury Is Still Out.

Tilsotolimod, an oligodeoxynucleotide TLR9 agonist, administered intratumorally, has been clinically evaluated. This compound has demonstrated the ability to induce changes within the tumor microenvironment, to convert noninflamed cold tumors into inflamed hot tumors, with the hope that these tumors will be more responsive to immune checkpoint blockade. See related article by Babiker et al., p. 5079.

Toll-like receptor 9 agonists and combination therapies: strategies to modulate the tumour immune microenvironment for systemic anti-tumour immunity.

Over the past decade, tremendous progress has taken place in tumour immunotherapy, relying on the fast development of combination therapy strategies that target multiple immunosuppressive signaling pathways in the immune system of cancer patients to achieve a high response rate in clinical practice. Toll-like receptor 9 (TLR9) agonists have been extensively investigated as therapeutics in monotherapy or combination therapies for the treatment of cancer, infectious diseases and allergies. TLR9 agonists monotherapy shows limited efficacy in cancer patients; whereas, in combination with other therapies including antigen vaccines, radiotherapies, chemotherapies and immunotherapies exhibit great potential. Synthetic unmethylated CpG oligodeoxynucleotide (ODN), a commonly used agonist for TLR9, stimulate various antigen-presenting cells in the tumour microenvironment, which can initiate innate and adaptive immune responses. Novel combination therapy approaches, which co-deliver immunostimulatory CpG-ODN with other therapeutics, have been tested in animal models and early human clinical trials to induce anti-tumour immune responses. In this review, we describe the basic understanding of TLR9 signaling pathway; the delivery methods in most studies; discuss the key challenges of each of the above mentioned TLR9 agonist-based combination immunotherapies and provide an overview of the ongoing clinical trial results from CpG-ODN based combination therapies in cancer patients.

Application of Transcription Factor Decoy Oligodeoxynucleotides (ODNs) for Cancer Therapy.

As a method of gene therapy, application of decoy oligodeoxynucleotides (ODNs) could interfere at the pretranscription level, by blocking the transcription factors, and inhibiting their attachment to the corresponding sequences in genomic DNA. Some of the transcription factors including MYC, OCT4, SOX2, STAT3, and NANOG are associated with the stemness properties of cancer cells, and suppressing them could interfere with cellular differentiation, which synergizes the efficiency of other anticancer therapies. The use of decoy ODNs has shown to be an effective measure against various malignancies, and it has shown to have a synergic effect when it is used along with the other cancer therapy methods. Emergence of modern nanocarriers has shown to further improve the outcome of using decoy ODNs against some cancers, and it has the potential of being used for clinical applications. In this chapter, it was aimed to provide a glance of this method for cancer therapy.

Introduction of the T315I gatekeeper mutation of BCR/ABL1 into a Philadelphia chromosome-positive lymphoid leukemia cell line using the CRISPR/Cas9 system.

Imatinib and second-generation tyrosine kinase inhibitors (TKIs) have dramatically improved the prognosis of Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL). However, overcoming TKI resistance due to the T315I gatekeeper mutation of BCR/ABL1 is crucial for further improving the prognosis. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is appropriate for establishing a human model of Ph+ ALL with the T315I mutation, because it can induce specific mutations via homologous recombination (HR) repair in cells with intact endogenous HR pathway. Here we used CRISPR/Cas9 to introduce the T315I mutation into the Ph+ lymphoid leukemia cell line KOPN55bi, which appeared to have an active HR pathway based on its resistance to a poly (ADP-Ribose) polymerase-1 inhibitor. Single-guide RNA targeting at codon 315 and single-strand oligodeoxynucleotide containing ACT to ATT nucleotide transition at codon 315 were electroporated with recombinant Cas9 protein. Dasatinib-resistant sublines were obtained after one-month selection with the therapeutic concentration of dasatinib, leading to T315I mutation acquisition through HR. T315I-acquired sublines were highly resistant to imatinib and second-generation TKIs but moderately sensitive to the therapeutic concentration of ponatinib. This authentic human model is helpful for developing new therapeutic strategies overcoming TKI resistance in Ph+ ALL due to T315I mutation.

Synergy of a STING agonist and an IL-2 superkine in cancer immunotherapy against MHC I-deficient and MHC I+ tumors.

Cyclic dinucleotides (CDN) and Toll-like receptor (TLR) ligands mobilize antitumor responses by natural killer (NK) cells and T cells, potentially serving as complementary therapies to immune checkpoint therapy. In the clinic thus far, however, CDN therapy targeting stimulator of interferon genes (STING) protein has yielded mixed results, perhaps because it initiates responses potently but does not provide signals to sustain activation and proliferation of activated cytotoxic lymphocytes. To improve efficacy, we combined CDN with a half life-extended interleukin-2 (IL-2) superkine, H9-MSA (mouse serum albumin). CDN/H9-MSA therapy induced dramatic long-term remissions of the most difficult to treat major histocompatibility complex class I (MHC I)­deficient and MHC I+ tumor transplant models. H9-MSA combined with CpG oligonucleotide also induced potent responses. Mechanistically, tumor elimination required CD8 T cells and not NK cells in the case of MHC I+ tumors and NK cells but not CD8 T cells in the case of MHC-deficient tumors. Furthermore, combination therapy resulted in more prolonged and more intense NK cell activation, cytotoxicity, and expression of cytotoxic effector molecules in comparison with monotherapy. Remarkably, in a primary autochthonous sarcoma model that is refractory to PD-1 checkpoint therapy, the combination of CDN/H9-MSA with checkpoint therapy yielded long-term remissions in the majority of the animals, mediated by T cells and NK cells. This combination therapy has the potential to activate responses in tumors resistant to current therapies and prevent MHC I loss accompanying acquired resistance of tumors to checkpoint therapy.

Review article: emerging drug therapies in inflammatory bowel disease.

BACKGROUND: The landscape of inflammatory bowel disease (IBD) treatment is rapidly expanding with the development of new therapeutic options. AIM: To review the mechanisms of action and the available clinical trial data on emerging drug therapies for IBD. METHODS: Pubmed, Medline and Cochrane databases were queried up to July 2021 using keywords "inflammatory bowel disease," "IBD," "Crohn's disease," "ulcerative colitis" and "trial," "phase" and "study." In addition, we manually reviewed the grey literature including clinical trial registries and abstracts from major gastroenterology conferences in 2020 and 2021 to include pertinent information. RESULTS: In ulcerative colitis (UC), phase 2b and/or phase 3 studies met primary endpoints for S1P receptor agonists (estrasimod, ozanimod), anti-IL-23 agent (mirikizumab), anti-lymphocyte trafficking agents (ontamalimab, subcutaneous vedolizumab), JAK inhibitors (upadacitinib, filgotinib) and TLR9 agonist (cobitolimod). In Crohn's disease (CD), anti-IL-23 agents (risankizumab, mirikizumab, guselkumab), JAK inhibitors (upadacitinib, filgotinib) and anti-lymphocyte trafficking agents (ontamalimab, etrolizumab) met primary endpoints in randomised controlled clinical trials. CONCLUSION: Several new IBD drug therapies have positive efficacy and safety data in early clinical trials, and there are several drugs in the therapeutic pipeline. As more treatments for CD and UC are approved for clinical use, research to assess predictors of response to therapy and head-to-head trials is needed to inform providers on how to best position therapeutic options for patients with IBD.

Efficacy of the adjuvanted subunit protein COVID-19 vaccine, SCB-2019: a phase 2 and 3 multicentre, double-blind, randomised, placebo-controlled trial.

BACKGROUND: A range of safe and effective vaccines against SARS CoV 2 are needed to address the COVID 19 pandemic. We aimed to assess the safety and efficacy of the COVID-19 vaccine SCB-2019. METHODS: This ongoing phase 2 and 3 double-blind, placebo-controlled trial was done in adults aged 18 years and older who were in good health or with a stable chronic health condition, at 31 sites in five countries (Belgium, Brazil, Colombia, Philippines, and South Africa). The participants were randomly assigned 1:1 using a centralised internet randomisation system to receive two 0·5 mL intramuscular doses of SCB-2019 (30 µg, adjuvanted with 1·50 mg CpG-1018 and 0·75 mg alum) or placebo (0·9% sodium chloride for injection supplied in 10 mL ampoules) 21 days apart. All study staff and participants were masked, but vaccine administrators were not. Primary endpoints were vaccine efficacy, measured by RT-PCR-confirmed COVID-19 of any severity with onset from 14 days after the second dose in baseline SARS-CoV-2 seronegative participants (the per-protocol population), and the safety and solicited local and systemic adverse events in the phase 2 subset. This study is registered on EudraCT (2020-004272-17) and ClinicalTrials.gov (NCT04672395). FINDINGS: 30 174 participants were enrolled from March 24, 2021, until the cutoff date of Aug 10, 2021, of whom 30 128 received their first assigned vaccine (n=15 064) or a placebo injection (n=15 064). The per-protocol population consisted of 12 355 baseline SARS-CoV-2-naive participants (6251 vaccinees and 6104 placebo recipients). Most exclusions (13 389 [44·4%]) were because of seropositivity at baseline. There were 207 confirmed per-protocol cases of COVID-19 at 14 days after the second dose, 52 vaccinees versus 155 placebo recipients, and an overall vaccine efficacy against any severity COVID-19 of 67·2% (95·72% CI 54·3-76·8), 83·7% (97·86% CI 55·9-95·4) against moderate-to-severe COVID-19, and 100% (97·86% CI 25·3-100·0) against severe COVID-19. All COVID-19 cases were due to virus variants; vaccine efficacy against any severity COVID-19 due to the three predominant variants was 78·7% (95% CI 57·3-90·4) for delta, 91·8% (44·9-99·8) for gamma, and 58·6% (13·3-81·5) for mu. No safety issues emerged in the follow-up period for the efficacy analysis (median of 82 days [IQR 63-103]). The vaccine elicited higher rates of mainly mild-to-moderate injection site pain than the placebo after the first (35·7% [287 of 803] vs 10·3% [81 of 786]) and second (26·9% [189 of 702] vs 7·4% [52 of 699]) doses, but the rates of other solicited local and systemic adverse events were similar between the groups. INTERPRETATION: Two doses of SCB-2019 vaccine plus CpG and alum provides notable protection against the entire severity spectrum of COVID-19 caused by circulating SAR-CoV-2 viruses, including the predominating delta variant. FUNDING: Clover Biopharmaceuticals and the Coalition for Epidemic Preparedness Innovations.

Nuclear factor-kappa B decoy oligodeoxynucleotide-loaded poly lactic-co-glycolic acid nanospheres promote periodontal tissue healing after tooth replantation in rats.

BACKGROUND: Excessive inflammation in the periodontal tissue after tooth replantation can lead to inflammatory root resorption and interrupt periodontal tissue regeneration. We tested the hypothesis that nuclear factor-κB decoy oligodeoxynucleotide-loaded poly lactic-co-glycolic acid nanospheres (NF-PLGA) inhibit excessive inflammation and promote healing of periodontal tissue after replantation in rats. METHODS: The upper right incisors of rats were extracted, immersed in different specific solutions, and replanted. The rats were euthanized at 7, 14, and 28 days after replantation. Morphological evaluation with micro-CT and histological assessment with hematoxylin and eosin and tartrate-resistant acid phosphatase (TRAP) staining was performed. Additionally, we examined the expression of interleukin (IL)-1ß, IL-6, transforming growth factor-ß1 (TGF-ß1), and fibroblast growth factor-2 (FGF-2) in the periodontal ligament (PDL) by performing immunohistological assessment. RESULTS: The NF-PLGA group showed significantly greater dental root thickness than the other experimental groups. Root resorption was not observed after the application of NF-PLGA on day 7. The application of NF-PLGA also resulted in a significantly lower number of TRAP-positive osteoclasts on days 7 and 14 after replantation. Significantly lower expression of IL-1ß and IL-6 and higher expression of TGF-ß1 and FGF-2 were observed under the application of NF-PLGA in the PDL. CONCLUSIONS: NF-PLGA promoted the healing process by inhibiting the initial excessive inflammatory response in the PDL, preventing root resorption, and promoting periodontal tissue regeneration. The findings also suggested that the PLGA nanospheres-mediated transfection of the decoy oligodeoxynucleotides can be useful for the clinical application of replanted tooth root surfaces.

Radiation Augments the Local Anti-Tumor Effect of In Situ Vaccine With CpG-Oligodeoxynucleotides and Anti-OX40 in Immunologically Cold Tumor Models.

Introduction: Combining CpG oligodeoxynucleotides with anti-OX40 agonist antibody (CpG+OX40) is able to generate an effective in situ vaccine in some tumor models, including the A20 lymphoma model. Immunologically "cold" tumors, which are typically less responsive to immunotherapy, are characterized by few tumor infiltrating lymphocytes (TILs), low mutation burden, and limited neoantigen expression. Radiation therapy (RT) can change the tumor microenvironment (TME) of an immunologically "cold" tumor. This study investigated the effect of combining RT with the in situ vaccine CpG+OX40 in immunologically "cold" tumor models. Methods: Mice bearing flank tumors (A20 lymphoma, B78 melanoma or 4T1 breast cancer) were treated with combinations of local RT, CpG, and/or OX40, and response to treatment was monitored. Flow cytometry and quantitative polymerase chain reaction (qPCR) experiments were conducted to study differences in the TME, secondary lymphoid organs, and immune activation after treatment. Results: An in situ vaccine regimen of CpG+OX40, which was effective in the A20 model, did not significantly improve tumor response or survival in the "cold" B78 and 4T1 models, as tested here. In both models, treatment with RT prior to CpG+OX40 enabled a local response to this in situ vaccine, significantly improving the anti-tumor response and survival compared to RT alone or CpG+OX40 alone. RT increased OX40 expression on tumor infiltrating CD4+ non-regulatory T cells. RT+CpG+OX40 increased the ratio of tumor-infiltrating effector T cells to T regulatory cells and significantly increased CD4+ and CD8+ T cell activation in the tumor draining lymph node (TDLN) and spleen. Conclusion: RT significantly improves the local anti-tumor effect of the in situ vaccine CpG+OX40 in immunologically "cold", solid, murine tumor models where RT or CpG+OX40 alone fail to stimulate tumor regression.

Aptamer-PROTAC Conjugates (APCs) for Tumor-Specific Targeting in Breast Cancer.

Development of proteolysis targeting chimeras (PROTACs) is emerging as a promising strategy for targeted protein degradation. However, the drug development using the heterobifunctional PROTAC molecules is generally limited by poor membrane permeability, low in vivo efficacy and indiscriminate distribution. Herein an aptamer-PROTAC conjugation approach was developed as a novel strategy to improve the tumor-specific targeting ability and in vivo antitumor potency of conventional PROTACs. As proof of concept, the first aptamer-PROTAC conjugate (APC) was designed by conjugating a BET-targeting PROTAC to the nucleic acid aptamer AS1411 (AS) via a cleavable linker. Compared with the unmodified BET PROTAC, the designed molecule (APR) showed improved tumor targeting ability in a MCF-7 xenograft model, leading to enhanced in vivo BET degradation and antitumor potency and decreased toxicity. Thus, the APC strategy may pave the way for the design of tumor-specific targeting PROTACs and have broad applications in the development of PROTAC-based drugs.

Cell surface nucleolin as active bait for nanomedicine in cancer therapy: a promising option.

Conventional chemotherapy used against cancer is mostly limited due to their non-targeted nature, affecting normal tissue and causing undesirable toxic effects to the affected tissue. With the aim of improving these treatments both therapeutically and in terms of their safety, numerous studies are currently being carried out using nanoparticles (NPs) as a vector combining tumor targeting and carrying therapeutic tools. In this context, it appears that nucleolin, a molecule over-expressed on the surface of tumor cells, is an interesting therapeutic target. Several ligands, antagonists of nucleolin of various origins, such as AS1411, the F3 peptide and the multivalent pseudopeptide N6L have been developed and studied as therapeutic tools against cancer. Over the last ten years or so, numerous studies have been published demonstrating that these antagonists can be used as tumor targeting agents with NPs from various origins. Focusing on nucleolin ligands, the aim of this article is to review the literature recently published or under experimentation in our research team to evaluate the efficacy and future development of these tools as anti-tumor agents.