After infection with the varicella-zoster virus (VZV), the virus becomes latent in the sensory ganglia. Immune senescence may lead to its reactivation, resulting in herpes zoster (HZ). The limited immunogenicity of current vaccines in elderly populations remains a significant challenge for prevention and control. This study investigated the immune-enhancing effects of the novel compound adjuvant BC02 on a recombinant VZV glycoprotein E (gE) subunit vaccine in a serum-positive elderly mouse model. A seropositive state was simulated through pre-immunization with the Oka strain of VZV, and the impacts of vaccines with various adjuvant formulations on humoral and cellular immunity in aged mice were systematically compared. Results demonstrated that the number of gE-specific IFN-γ- and IL-2-secreting cells induced by the BC02-adjuvanted vaccine (gE+BC02-1) increased 11.8- and 5.7-fold compared to the single-adjuvant group, significantly enhancing the multifunctionality of CD4+ T cells. The neutralizing antibody titer reached 1:122, comparable to that of the commercial vaccine Shingrix®, while the ratio of memory T/B cells was markedly higher than in the control group. Cross-age group experiments revealed that BC02 could overcome the limitations imposed by immune senescence in elderly models, inducing a balanced Th1/Th2 response and long-term immune memory comparable to that observed in younger groups (antibody titers maintained for ≥8 months). This study confirmed that the BC02 adjuvant synergistically activates innate and adaptive immunity, significantly enhancing the immune efficacy of the gE vaccine in serum-positive elderly individuals, thereby providing an potential strategy for optimizing herpes zoster vaccines for the elderly population.

Since 2019, there has been a growing focus on mRNA vaccines for infectious disease prevention, particularly following the emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). mRNA vaccines offer advantages such as rapid production and the ability to induce robust cellular and antibody responses, which are essential for combating infections that require cell-mediated immunity, including Tuberculosis (TB). This review explores recent progress in TB mRNA vaccines and addresses several key areas: (1) the urgent need for new TB vaccines; (2) current advancements in TB vaccine development, and the advantages and challenges of mRNA technology; (3) the design and characteristics of TB mRNA vaccines; (4) the immunological mechanisms of TB mRNA vaccines; (5) manufacturing processes for TB mRNA vaccines; and (6) safety and regulatory considerations. This interdisciplinary review aims to provide insights for researchers working to address critical questions in TB mRNA vaccine development.

Messenger RNA (mRNA) vaccines have revolutionized infectious disease prevention and cancer immunotherapy due to their rapid development, potent immunogenicity, and flexible design. Central to the clinical success of mRNA vaccines, lipid nanoparticles (LNPs) function as efficient, nonviral delivery systems capable of protecting mRNA and facilitating its uptake by target cells. Recent advances have demonstrated that LNP-formulated mRNA vaccines and therapeutics elicit robust immune responses and confer effective protection against a broad spectrum of pathogens, including viruses and bacteria. Moreover, LNP-based therapies have shown promising therapeutic efficacy in various cancers and rare diseases, as evidenced by both preclinical models and clinical trials. This review provides a comprehensive overview of the key components, structural features, and preparation technologies of LNPs. It further discusses ongoing challenges in LNP design, such as delivery efficiency, tissue targeting, and safety, and proposes rational strategies to address these limitations. Additionally, recent progress in the analytical methods used to characterize the critical quality attributes of LNPs is highlighted. This review aims to guide the rational design of next-generation LNPs and to support the broader application of mRNA-based vaccines and therapeutics.

Tuberculosis (TB) remains one of the most significant challenges to global public health. Vaccine development is a critical strategy for the prevention and control of TB. However, evaluating the protective efficacy of TB vaccines faces numerous challenges, particularly in the selection of animal models and bacterial strains. Variations in animal models, challenge strains, challenge routes, and doses can significantly impact the outcomes of preclinical evaluations. This article highlights the importance of standardizing preclinical evaluation models, summarizes the animal models and challenge strains used in novel TB vaccine candidates, efficacy studies, and discusses the advantages and limitations of commonly used animal models in TB vaccine research. It also points out the differential performance of various animal models in simulating protection and pathology. Given the current limitations of using a narrow range of challenge strains and the lack of standardized infection routes and doses, this article calls for the establishment of more standardized challenge strains and the development of standardized evaluation models to improve the reliability and generalizability of new TB vaccine efficacy assessments.

The increasing challenge of drug-resistant tuberculosis (TB) calls for the development of innovative therapeutic strategies, highlighting the potential of adjunctive immunotherapies that are both cost-effective and safe. Host-directed therapy (HDT) using immunomodulators shows promise in enhancing treatment efficacy by modulating immune responses, thereby shortening the duration of therapy and reducing drug resistance risks. This study investigated the immunomodulatory potential of combining Heat-killed Bacillus Calmette-Guérin (hBCG) with a Squalene-based oil-in-Water Emulsion (SWE) adjuvant against TB. The therapeutic efficacy of the hBCG-SWE regimen was assessed in a guinea pig model infected with Mycobacterium tuberculosis (M. tb). Furthermore, the impact of hBCG-SWE on TNF-α and MCP-1 production was evaluated in RAW264.7 macrophages, examining the role of TLR2/4 and MyD88 signaling pathways using ELISA, both with and without specific inhibitors. Our findings revealed that hBCG-SWE significantly enhanced TNF-α and MCP-1 production compared to hBCG alone, indicating activation through TLR2/4 and MyD88-dependent pathways. In guinea pigs, hBCG-SWE administration led to notable reductions in lung pathology and spleen bacterial loads versus control groups. These results highlight the capacity of hBCG-SWE to boost innate immunity and provide robust protection against M. tb. Future research should focus on evaluating the ability of hBCG-SWE to shorten conventional chemotherapy and exploring ways to amplify its immunomodulatory efficacy through advanced formulation techniques.

The blockade of programmed death-ligand 1 (PD-L1) pathway has been clinically used in cancer immunotherapy, while its effects on infectious diseases remain elusive. Roles of PD-L1 signaling in the macrophage-mediated innate immune defense against M.tb is unclear. In this study, the outcomes of tuberculosis (TB) in wild-type (WT) mice treated with anti-PD-1/PD-L1 therapy and macrophage-specific Pdl1-knockout (Pdl1ΔΜΦ) mice were compared. Treatment with anti-PD-L1 or anti-PD-1 benefited protection against M.tb infection in WT mice, while Pdl1ΔΜΦ mice exhibited the increased susceptibility to M.tb infection. Mechanistically, the absence of PD-L1 signaling impaired M.tb killing by macrophages. Furthermore, elevated STAT3 activation was found in PD-L1-deficient macrophages, leading to increased interleukin (IL)-6 production and reduced inducible nitric oxide synthase (iNOS) expression. Inhibiting STAT3 phosphorylation partially impeded the increase in IL-6 production and restored iNOS expression in these PD-L1-deficient cells. These findings provide valuable insights into the complexity and mechanisms underlying anti-PD-L1 therapy in the context of tuberculosis.

Novel adjuvants and innovative combinations of adjuvants (Adjuvant Systems) have facilitated the development of enhanced and new vaccines against re-emerging and challenging pathogenic microorganisms. Nonetheless, the efficacy of adjuvants is influenced by various factors, and the same adjuvant may generate entirely different immune responses when paired with different antigens. Herein, we combined the MPXV-B6R antigen with BC02, a novel adjuvant with proprietary technology, to assess its capability to induce both cellular and humoral immunity in mouse models. Mice received two intramuscular injections of B6R-BC02, which resulted in the production of MPXV-specific IgG, IgG1, and IgG2a antibodies. Additionally, it elicited strong MPXV-specific Th1-oriented cellular immunity and persistent effector memory B-cell responses. The advantages of BC02 were further validated, including rapid initiation of the immune response, robust recall memory, and sustained immune response induction. Although the potential of immunized mice to produce serum-neutralizing antibodies against the vaccinia virus requires further improvement, the exceptional performance of BC02 as an adjuvant for the MPXV-B6R antigen has been consistently demonstrated.

To achieve maximum efficacy, vaccines, such as subunit, recombinant, and conjugate vaccines, necessitate the incorporation of immunostimulators/adjuvants. Adjuvants play a vital role in bolstering and extending the strength of the immune response while also influencing its type. As antigen and adjuvant formulations become more intricate, it becomes imperative to establish a well-characterized and robust formulation to ensure consistent and reproducible outcomes in preclinical and clinical studies. In the present study, an HPV bivalent vaccine was developed using a BC02 adjuvant in conjunction with HPV 16 and 18 L1 VLP antigens produced from an E. coli expression system. The study involved evaluating the adjuvant formulation and in vivo immunogenicity in mice. Remarkably, a medium-dose of BCG-CpG-DNA combined with a low-dose of aluminum hydroxide substantially enhanced the immunogenicity of HPV16 and 18 VLPs, resulting in improved cellular and humoral immune responses.

While purified protein derivative (PPD) is commonly used as skin diagnostic reagent for tuberculosis (TB) infection, it cannot distinguish effectively Bacillus Calmette–Guérin (BCG) vaccination from Mycobacterium tuberculosis (MTB) complex and nontuberculous mycobacteria infection. The new skin reagent ESAT6-CFP10 (EC) has favorable sensitivity and specificity, which can overcome limitations associated with PPD. At present, EC skin test reactions are mainly characterized by erythema, while PPD mainly causes induration. We conducted a comparative study on the potential differences between EC-induced erythema and PPD-induced induration using a guinea pig model. The size of EC-dependent erythema was similar to that of PPD-induced induration, and an inflammatory response characterized by the infiltration of monocytes, macrophages and lymphocytes, as well as tissue damage, appeared at the injection site. The lymphocytes included CD4+ T and CD8+ T cells, which released IFN-γ as the main cytokine. Both EC erythema and PPD induration could lead to increased levels of acute-phase proteins, and the differential pathways were similar, thus indicating that the main induced immune pathways were similar. The above results indicated that erythema produced by EC could generate the main delayed-type hypersensitivity (DTH) response characteristic of PPD induration, thereby suggesting that erythema might also have a certain diagnostic significance and provide a possible theoretical basis for its use as a diagnostic indicator for detecting MTB infection.

A latent tuberculosis infection (LTBI) is a major source of active tuberculosis, and addressing an LTBI is crucial for the elimination of tuberculosis. The treatment of tuberculosis often requires a 6-month course of multidrug therapy, and for drug-resistant tuberculosis, a longer course of multidrug therapy is needed, which has many drawbacks. At present, vaccines are proposed as an adjunct to chemotherapy to protect populations with an LTBI and delay its recurrence. In this study, we analyzed the protective effect of a novel subunit vaccine, AEC/BC02, in a guinea pig latent infection model. Through the optimization of different chemotherapy durations and immunization times, it was found that 4 weeks of administration of isoniazid–rifampin tablets combined with three or six injections of the vaccine could significantly reduce the gross pathological score and bacterial load in organs and improve the pathological lesions. This treatment regimen had a better protective effect than the other administration methods. Furthermore, no drug resistance of Mycobacterium tuberculosis was detected after 2 or 4 weeks of administration of the isoniazid–rifampin tablets, indicating a low risk of developing drug-resistant bacteria during short-term chemotherapy. The above results provided the foundation for an AEC/BC02 clinical protocol.

目的 研究宿主铁元素吸收相关转铁蛋白受体1(transferrin receptor,TfR1)对结核分枝杆菌(Mycobacterium tubercu-losis,M.tb)胞内生存的影响,探讨其成为抗结核新靶点的可能.方法 采用免疫组化和蛋白质免疫印迹技术对不同剂量铁过载小鼠以及M.tb感染小鼠肺组织切片和肺部组织研磨液上清TfR1表达丰度进行检测,提取肺组织总RNA并进行TfR1基因qPCR验证.siRNA技术构建TfR1--RAW 264.7巨噬细胞系,以FITC标记的耻垢分枝杆菌(FITC-Mycobacterium smegmatis,FITC-M.smegmatis)和M.smegmatis进行巨噬细胞吞噬和杀伤功能实验,并通过流式细胞术和细菌培养菌落计数方法进行检测分析.结果 低剂量、中剂量和高剂量铁剂小鼠肺组织TfR1表达水平与阴性对照小鼠差异无统计学意义.M.tb感染后,铁过载M.tb感染小鼠和单纯M.tb感染小鼠肺组织TfR1表达均显著上调,且前者TfR1的表达丰度显著高于后者.下调RAW 264.7巨噬细胞TfR1表达水平后,TfR1-/-与TfR1+/+RAW 264.7巨噬细胞在1、2和4h吞噬FITC-M.smegmatis数量组间差异无统计学意义.M.smegmatis感染后24、48和72h后,TfR1-/-RAW 264.7巨噬细胞杀伤功能则明显优于TfR1+/+RAW 264.7巨噬细胞,胞内残余活M.smegmatis数量显著低于未转染的TfR1+/+正常细胞.结论 TfR1参与了M.tb的侵染,并可能与M.tb获取铁元素以利其在胞内生存有关;TfR1的下调不影响M.tb的入侵,但抑制其在巨噬细胞内的增殖.

Herpes zoster (HZ) is one of two distinct syndromes caused by Varicella-zoster virus (VZV). A primary infection with VZV causes varicella in susceptible young children. After resolution of the primary infection, VZV establishes a lifelong latency within the cranial or dorsal root ganglia. With increasing age, family history of shingles, immunosuppression or other risk factors, there is a decline in the virus-specific T-cell-mediated immune (CMI) response which allows reactivation of latent VZV in the root ganglia resulting in HZ. There are currently two vaccines that have been approved to prevent HZ and postherpetic neuralgia (PHN) but one is a live attenuated vaccine, the protective effect of which is considered to decrease significantly with the age of the recipient. However, a recombinant subunit vaccine may provide targeted VZV-specific cellular and humoral immunity, giving it a more potent and longer-lasting protective effect against HZ. The current study reports the development of a novel adjuvant, BC02 (BCG CpG DNA compound adjuvants system 02), composed of Al(OH)3 inorganic salt adjuvant and BC01 (BCG CpG DNA compound adjuvants system 01), a Toll-like receptor 9 (TLR9) agonist. Immunogenicity and compatibility with recombinant VZV glycoprotein E (gE) in mice were studied. The BC02-adjuvanted gE experimental vaccine was highly effective in eliciting both humoral and cellular immune responses to the recombinant gE glycoprotein and VZV-Oka in a mouse model. The efficient production and long-term persistence of gE and VZV-Oka-specific IFN-γ, IL-2-specific T cells and memory B cells in the early (1W), middle (7W), middle-late (15W), and final (27W) immune stages were established. Results of fluorescent antibody to membrane antigen (FAMA) and serum antibody plaque reduction tests also showed that the BC02 adjuvanted-gE experimental vaccine induced mice to secrete neutralizing antibodies against clinically isolated VZV strains. In combination, the current data suggest that the BC02 compound adjuvant offers a strategy to induce an appropriately strong cellular and humoral immunity against the VZV gE protein subunit to improve vaccine efficacy.

为实现到2050年在全球范围内根除结核病(Tuberculosis,TB)这一宏伟目标,迫切需要开发新的TB疫苗和疫苗接种策略.虽然卡介苗(Bacille Calmette-Guérin,BCG)在保护儿童免受粟粒性结核和结核性脑膜炎等方面获得了成功,但其对成人肺结核(Pulmonary tuberculosis,PTB)的保护效果仍然存在较大差异(0～80％).目前,有包括第一代疫苗和第二代疫苗在内的几十种新型TB疫苗已处于临床前或临床研究的不同阶段,而作为第三代疫苗的核酸疫苗随着医药技术的发展也逐渐走入人们的视野.本文主要综述了 TBDNA疫苗的免疫学机制、研究现状、免疫增强策略和TBDNA疫苗存在的问题与应用前景,以期为开发更安全、更高效的新型TB DNA疫苗提供新的思路和视角.

目的 初步探讨新型生物佐剂BC01(BCG CpG DNA compound adjuvant system 01)对机体的免疫激活作用.方法 将C57BL/6小鼠随机分为4组并分别肌肉注射75 μg新型生物佐剂BC01,实验组于免疫后6、12和24 h解剖取脾脏,对照组于注射BC01后0h解剖取脾脏.提取各组小鼠总RNA并进行全基因转录组分析.基因芯片数据经归一化处理后进行基因集富集分析(gene set enrichment analysis,GSEA).同时,采用实时定量PCR法对GSEA中 GO(gene ontology,GO)基因集分组归类中生物过程(biological process,BP)、细胞成分(cellular component,CC)和分子功能(molecular function,MF)相关上调基因进行验证分析.结果 小鼠全基因转录组芯片数据经GSEA分析后发现,BC01佐剂具有活化免疫细胞,激活固有免疫和适应性免疫应答,调节炎症反应、活化免疫细胞迁移与趋化,激活免疫细胞吞噬功能以及促分裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)和细胞外调节蛋白激酶1/2(extracellular-regulated kinase 1/2,ERK1/2)信号级联反应作用.结论 新型生物佐剂BC01经肌肉注射24 h后具有较强的机体免疫激活作用.

The BCG vaccine is prepared from a weakened strain of Mycobacterium bovis (M. bovis), a bacterium closely related to Mycobacterium tuberculosis (MTB), which causes tuberculosis (TB). The vaccine was developed over 13 years, from 1908 to 1921, in the French Institut Pasteur by Léon Charles Albert Calmette and Jean-Marie Camille Guérin, who named the product Bacillus Calmette–Guérin (BCG). BCG, the only licensed vaccine currently available to prevent TB, is given to infants at high risk of TB shortly after birth to protect infants and young children from pulmonary, meningeal, and disseminated TB. The BCG vaccine, one of the safest and most widely used live attenuated vaccines in the world, recently celebrated its 100th anniversary (from 1921 to 2021); its record of use in preventing TB in China is also approaching 100 years. In 2022, a new century of BCG vaccine immunization will begin. In this article, we briefly review the history of BCG vaccine use in China, describe its current status, and offer a preliminary outlook on the future of the vaccine, to provide BCG researchers with a clearer understanding of its use in China.

Angiotensin-converting enzyme 2 (ACE2) is required for the cellular entry of the severe acute respiratory syndrome coronavirus 2. ACE2, via the Ang-(1-7)-Mas-R axis, is part of the antihypertensive and cardioprotective effects of the renin-angiotensin system. We studied hospitalized COVID-19 patients with hypertension and hypertensive human(h) ACE2 transgenic mice to determine the outcome of COVID-19 with or without AT1 receptor (AT1R) blocker treatment. The severity of the illness and the levels of serum cardiac biomarkers (CK, CK-BM, cTnI), as well as the inflammation markers (IL-1, IL-6, CRP), were lesser in hypertensive COVID-19 patients treated with AT1R blockers than those treated with other antihypertensive drugs. Hypertensive hACE2 transgenic mice, pretreated with AT1R blocker, had increased ACE2 expression and SARS-CoV-2 in the kidney and heart, 1 day post-infection. We conclude that those hypertensive patients treated with AT1R blocker may be at higher risk for SARS-CoV-2 infection. However, AT1R blockers had no effect on the severity of the illness but instead may have protected COVID-19 patients from heart injury, via the ACE2-angiotensin1-7-Mas receptor axis.

目的 探讨复合佐剂BC02(BCG CpG DNA combination adjuvant system 02)组成成分对机体固有免疫应答的协同增强作用.方法 将C57BL/6小鼠随机分为4组,分别肌肉注射Al(OH)3佐剂、BC01佐剂和BC02复合佐剂及PBS溶液(对照).免疫后6h,将各组部分小鼠解剖取腹股沟淋巴结进行全基因组转录谱分析.免疫后24、48、72和96 h,将各组剩余小鼠解剖取右后肢内侧注射部位肌肉组织,经组织HE和免疫荧光染色,分析注射部位炎性细胞招募情况.同时,采用qPCR法对信号通路中富集的显著性差异表达基因(significantly differentially expressed genes,SDEG)进行验证分析.结果 SDEG数据经IPA(Ingenuity Pathway Analysis)软件进行GO富集和信号通路网络分析后发现,BC02复合佐剂组成成分具有协同增强免疫细胞活化网络激活、免疫细胞招募信号网络激活、巨噬细胞吞噬功能、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)信号通路网络激活和CCL2相互作用网络激活作用.BC02复合佐剂组成成分也具有协同增强佐剂在注射部位炎性细胞招募的能力.结论 BC02复合佐剂组成成分Al(OH)3无机盐佐剂及BC01生物佐剂具有协同增强机体固有免疫应答的作用.

Both adaptive and innate immunity responses are necessary for the efficient elimination of different pathogens. However, the magnitude, quality and desired type of immune response specific to the co-administered antigen is largely determined by adjuvants. BC02 (BCG CpG DNA compound adjuvants system 02) is a novel compound adjuvant with independent intellectual properties, which is composed of BCG CpG DNA biological adjuvant with Al(OH)3 inorganic salt adjuvant acting as a delivery system. Its safety and strong adjuvant efficacy have been effectively verified in preclinical and clinical trials (Phase Ib, ClinicalTrials.gov Identifier: NCT04239313 and Phase II, ClinicalTrials.gov Identifier: NCT05284812). In this study, we report the level of cell-mediated immunity (CMI) and humoral immune response induced by the BC02 novel adjuvant combined with different doses of varicella-zoster virus (VZV) glycoprotein E (gE) in a mouse model. In addition, we conducted preliminary in vitro experiments to explore the enhancement of RAW264.7 cell immune activity by BC02 adjuvanted-gE experimental vaccine to activate innate immune response. The results showed that the BC02-adjuvanted low, medium or high dose of gE were highly effective in eliciting both CMI and humoral immune responses to the immunized mice, respectively. The production of gE-specific IFN-γ and IL-2-specific T cells was established within 28 days after booster immunization. In particular, the effect of BC02-adjuvanted medium dose of gE has been shown to be more prominent. Meanwhile, fluorescent antibody to membrane antigen (FAMA) and serum antibody plaque reduction tests have also shown that the BC02 adjuvanted-medium dose of gE antigen could induce the secretion of neutralizing antibodies against clinically isolated VZV strains in mice. In addition, our findings have shown that 1/25 dose of gE+BC02 medium dose experimental vaccine can significantly induce the secretion of innate immune cytokines TNF-A, MCP-1, IL-6 and GM-CSF and up-regulate the costimulatory molecules CD40, CD80 and I-A/I-E on RAW264.7 cells; and it has also been activated to form M2 macrophages. At the same time, RAW264.7 cells were stimulated for 12 h, and their phagocytosis was significantly enhanced. Taken together, these results suggest that the BC02 compound adjuvant offers a strategy to induce an appropriate innate and adaptive immunity against the different doses of the VZV gE protein to improve subunit vaccine efficacy, and BC02 may be a promising adjuvant candidate for subunit HZ vaccines.

Bacillus Calmette–Guérin (BCG), the only vaccine proven to be effective against tuberculosis (TB), is the most commonly used vaccine globally. In addition to its effects on mycobacterial diseases, an increasing amount of epidemiological and experimental evidence accumulated since its introduction in 1921 has shown that BCG also exerts non-specific effects against a number of diseases, such as non-mycobacterial infections, allergies and certain malignancies. Recent Corona Virus Disease 2019 (COVID-19) outbreak has put BCG, a classic vaccine with significant non-specific protection, into the spotlight again. This literature review briefly covers the diverse facets of BCG vaccine, providing new perspectives in terms of specific and non-specific protection mechanisms of this old, multifaceted, and controversial vaccine.