Squalene-epoxidase-catalyzed 24(S),25-epoxycholesterol synthesis promotes trained-immunity-mediated antitumor activity.

The importance of trained immunity in antitumor immunity has been increasingly recognized, but the underlying metabolic regulation mechanisms remain incompletely understood. In this study, we find that squalene epoxidase (SQLE), a key enzyme in cholesterol synthesis, is required for ß-glucan-induced trained immunity in macrophages and ensuing antitumor activity. Unexpectedly, the shunt pathway, but not the classical cholesterol synthesis pathway, catalyzed by SQLE, is required for trained immunity induction. Specifically, 24(S),25-epoxycholesterol (24(S),25-EC), the shunt pathway metabolite, activates liver X receptor and increases chromatin accessibility to evoke innate immune memory. Meanwhile, SQLE-induced reactive oxygen species accumulation stabilizes hypoxia-inducible factor 1α protein for metabolic switching into glycolysis. Hence, our findings identify 24(S),25-EC as a key metabolite for trained immunity and provide important insights into how SQLE regulates trained-immunity-mediated antitumor activity.

Recent progress in mRNA cancer vaccines.

The research and development of messenger RNA (mRNA) cancer vaccines have gradually overcome numerous challenges through the application of personalized cancer antigens, structural optimization of mRNA, and the development of alternative RNA-based vectors and efficient targeted delivery vectors. Clinical trials are currently underway for various cancer vaccines that encode tumor-associated antigens (TAAs), tumor-specific antigens (TSAs), or immunomodulators. In this paper, we summarize the optimization of mRNA and the emergence of RNA-based expression vectors in cancer vaccines. We begin by reviewing the advancement and utilization of state-of-the-art targeted lipid nanoparticles (LNPs), followed by presenting the primary classifications and clinical applications of mRNA cancer vaccines. Collectively, mRNA vaccines are emerging as a central focus in cancer immunotherapy, offering the potential to address multiple challenges in cancer treatment, either as standalone therapies or in combination with current cancer treatments.

The advances of adjuvants in mRNA vaccines.

The remarkable success of messenger RNA (mRNA) vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has propelled the rapid development of this vaccination technology in recent years. Over the last three decades, numerous studies have shown the considerable potential of mRNA vaccines that elicit protective immune responses against pathogens or cancers in preclinical studies or clinical trials. These effective mRNA vaccines usually contain specific adjuvants to obtain the desired immune effect. Vaccine adjuvants traditionally are immunopotentiators that bind to pattern recognition receptors (PRRs) of innate immune cells to increase the magnitude or achieve qualitative alteration of immune responses, finally enhancing the efficacy of vaccines. Generally, adjuvants are necessary parts of competent vaccines. According to the existing literature, adjuvants in mRNA vaccines can be broadly classified into three categories: 1) RNA with self-adjuvant characteristics, 2) components of the delivery system, and 3) exogenous immunostimulants. This review summarizes the three types of adjuvants used in mRNA vaccines and provides a comprehensive understanding of molecular mechanisms by which adjuvants exert their functions in mRNA vaccines.

A Nanovaccine Based on Adjuvant Peptide FK-13 and l-Phenylalanine Poly(ester amide) Enhances CD8+ T Cell-Mediated Antitumor Immunity.

Cancer vaccines have shown promise as effective means of antitumor immunotherapy by inducing tumor antigen-specific T cell immunity. In this study, a novel peptide-based tumor nanovaccine that boosts antigen presentation and elicits effective antitumor immunity is developed. The adjuvant characteristics of an antimicrobial peptide-derived core peptide, FK-13, are investigated and used it to generate a fusion peptide named FK-33 with tumor antigen epitopes. l-phenylalanine-based poly(ester amide) (Phe-PEA), 8p4, is also identified as a competent delivery vehicle for the fusion peptide FK-33. Notably, the vaccination of 8p4 + FK-33 nanoparticles (8FNs) in vivo induces dendritic cell activation in the lymph nodes and elicits robust tumor antigen-specific CD8+ T cell response. The nanovaccine 8FNs demonstrate significant therapeutic and prophylactic efficacy against in situ tumor growth, effectively inhibit tumor metastasis, and significantly prolong the survival of tumor-bearing mice. Moreover, 8FNs can incorporate different tumor antigens and exhibit a synergistic therapeutic effect with antiprogrammed cell death protein 1 (PD-1) therapy. In summary, 8FNs represent a promising platform for personalized cancer vaccines and may serve as a potential combinational modality to improve current immunotherapy.

Advances in mRNA vaccines.

mRNA vaccines have been increasingly recognized as a powerful vaccine platform since the FDA approval of two COVID-19 mRNA vaccines, which demonstrated outstanding prevention efficacy as well as great safety profile. Notably, nucleoside modification and lipid nanoparticle-facilitated delivery has greatly improved the immunogenicity, stability, and translation efficiency of mRNA molecule. Here we review the recent progress in mRNA vaccine development, including nucleoside modification, in vitro synthesis and product purification, and lipid nanoparticle vectors for in vivo delivery and efficient translation. We also briefly introduce the clinical application of mRNA vaccine in preventing infectious diseases and treating inflammatory diseases including cancer.

Mevalonate Blockade in Cancer Cells Triggers CLEC9A+ Dendritic Cell-Mediated Antitumor Immunity.

Hyperactive mevalonate (MVA) metabolic activity is often observed in cancer cells, and blockade of this pathway inhibits tumor cell lipid synthesis and cell growth and enhances tumor immunogenicity. How tumor cell MVA metabolic blockade promotes antitumor immune responses, however, remains unclear. Here we show that inhibition of the MVA metabolic pathway in tumor cells elicits type 1 classical dendritic cells (cDC1)-mediated tumor recognition and antigen cross-presentation for antitumor immunity. Mechanistically, MVA blockade disrupted prenylation of the small GTPase Rac1 and induced cancer cell actin filament exposure, which was recognized by CLEC9A, a C-lectin receptor specifically expressed on cDC1s, in turn activating antitumor T cells. MVA pathway blockade or Rac1 knockdown in tumor cells induced CD8+ T-cell-mediated antitumor immunity in immunocompetent mice but not in Batf3 -/- mice lacking CLEC9A+ dendritic cells. These findings demonstrate tumor MVA metabolic blockade stimulates a cDC1 response through CLEC9A-mediated immune recognition of tumor cell cytoskeleton, illustrating a new immune surveillance mechanism by which dendritic cells monitor tumor metabolic dysregulation and providing insight into how MVA pathway inhibition may potentiate anticancer immunity. SIGNIFICANCE: These findings suggest that mevalonate blockade in cancer cells disrupts Rac1 prenylation to increase recognition and cross-presentation by conventional dendritic cells, suggesting this axis as a potential target for cancer immunotherapy.

Modulation of lactate-lysosome axis in dendritic cells by clotrimazole potentiates antitumor immunity.

BACKGROUND: Dendritic cells (DCs) play a critical role in antitumor immunity, but the therapeutic efficacy of DC-mediated cancer vaccine remains low, partly due to unsustainable DC function in tumor antigen presentation. Thus, identifying drugs that could enhance DC-based antitumor immunity and uncovering the underlying mechanism may provide new therapeutic options for cancer immunotherapy. METHODS: In vitro antigen presentation assay was used for DC-modulating drug screening. The function of DC and T cells was measured by flow cytometry, ELISA, or qPCR. B16, MC38, CT26 tumor models and C57BL/6, Balb/c, nude, and Batf3-/- mice were used to analyze the in vivo therapy efficacy and impact on tumor immune microenvironment by clotrimazole treatment. RESULTS: By screening a group of small molecule inhibitors and the US Food and Drug Administration (FDA)-approved drugs, we identified that clotrimazole, an antifungal drug, could promote DC-mediated antigen presentation and enhance T cell response. Mechanistically, clotrimazole acted on hexokinase 2 to regulate lactate metabolic production and enhanced the lysosome pathway and Chop expression in DCs subsequently induced DC maturation and T cell activation. Importantly, in vivo clotrimazole administration induced intratumor immune infiltration and inhibited tumor growth depending on both DCs and CD8+ T cells and potentiated the antitumor efficacy of anti-PD1 antibody. CONCLUSIONS: Our findings showed that clotrimazole could trigger DC activation via the lactate-lysosome axis to promote antigen cross-presentation and could be used as a potential combination therapy approach to improving the therapeutic efficacy of anti-PD1 immunotherapy.

miR-34a exerts as a key regulator in the dedifferentiation of osteosarcoma via PAI-1-Sox2 axis.

Osteosarcoma (OS) is a malignant bone cancer with severe chromosomal abnormalities and genetic aberrations. Our previous work reported the dedifferentiation of OS, which is related to poor prognosis. However, the molecular mechanism that regulates OS dedifferentiation is still a subject of exploration. Emerging evidence has suggested that microRNAs (miRNAs) are associated with the pathogenesis of OS and could potentially be developed for use as diagnostic biomarkers and therapeutic strategies. In the present study, we intended to illustrate the role of miR-34a in the dedifferentiation of OS. Upregulation of miR-34a was observed while OS cells were induced into stem-like phenotype. Notably, inhibition of miR-34a could promote the reprogramming transition of OS. Further exploration on the downstream network of miR-34a identified that blocking plasminogen activator inhibitor-1 (PAI-1) expression could restrain OS dedifferentiation into cancer stem-like cells by downregulating SRY-related-HMG box (Sox) 2. We also showed that Sox2 overexpression rescued the suppression phenotype driven by PAI-1 inhibition. Conversely, PAI-1 inhibitor (PAI-039) could suppress the upregulation of Sox2 expression caused by miR-34a inhibition. Be applying bone extracellular matrix (BEM)-OS models, we demonstrated the phenotypic heterogeneity of OS cells, consistent with a strong concordance between PAI-1 and Sox2 expression levels. Taken together, our findings proved miR-34a to be a bona fide suppressor involved in the regulation of OS dedifferentiation. Targeting miR-34a or its direct target PAI-1 could offer new strategies for OS treatment.

Estimation of the optimal ratio of standardized ileal digestible threonine to lysine for finishing barrows fed low crude protein diets.

Two experiments were conducted to determine the standardized ileal digestible (SID) lysine (Lys) requirement and the ideal SID threonine (Thr) to Lys ratio for finishing barrows. In Exp. 1, 120 barrows with an average body weight of 72.8±3.6 kg were allotted to one of six dietary treatments in a randomized complete block design conducted for 35 d. Each diet was fed to five pens of pigs containing four barrows. A normal crude protein (CP) diet providing 15.3% CP and 0.71% SID Lys and five low CP diets providing 12% CP with SID Lys concentrations of 0.51, 0.61, 0.71, 0.81 and 0.91% were formulated. Increasing the SID Lys content of the diet resulted in an increase in weight gain (linear effect p = 0.04 and quadratic effect p = 0.08) and an improvement in feed conversion ratio (FCR) (linear effect p = 0.02 and quadratic effect p = 0.02). For weight gain and FCR, the estimated SID Lys requirement of finishing barrows were 0.71 and 0.71% (linear broken-line analysis), 0.79 and 0.78% (quadratic analysis), respectively. Exp. 2 was a 26 d dose-response study using SID Thr to Lys ratios of 0.56, 0.61, 0.67, 0.72 and 0.77. A total of 138 barrows weighing 72.5±4.4 kg were randomly allotted to receive one of the five diets. All diets were formulated to contain 0.61% SID Lys (10.5% CP), which is slightly lower than the pig's requirement. Weight gain was quadratically (p = 0.03) affected by SID Thr to Lys ratio while FCR was linearly improved (p = 0.02). The SID Thr to Lys ratios for maximal weight gain and minimal FCR and serum urea nitrogen (SUN) were 0.67, 0.71 and 0.64 using a linear broken-line model and 0.68, 0.78 and 0.70 using a quadratic model, respectively. Based on the estimates obtained from the broken-line and quadratic analysis, we concluded that the dietary SID Lys requirement for both maximum weight gain and minimum FCR was 0.75%, and an optimum SID Thr to Lys ratio was 0.68 to maximize weight gain, 0.75 to optimize FCR and 0.67 to minimize SUN for finishing barrows.

Effects of the antimicrobial peptide cecropin AD on performance and intestinal health in weaned piglets challenged with Escherichia coli.

This study was conducted to determine the effects of the antimicrobial peptide cecropin on performance and intestinal health in piglets. Newly weaned barrows were randomly assigned to one of three treatments (n=8), including a corn-soybean basal diet or similar diets supplemented with antibiotics (100 mg/kg kitasamycin plus 800 mg/kg colistin sulfate) or 400 mg/kg cecropin AD. On day 13, all piglets were orally challenged with 10(9)CFU/mL of Escherichia coli K88. On day 19, all piglets were euthanized and sampled. Before challenge, piglets fed antibiotics had greater weight gain, feed efficiency, nitrogen and energy retention than the control (P<0.05). E. coli challenge decreased weight gain, feed intake and feed efficiency for the control piglets (P<0.05) but not for the antibiotic or cecropin AD treated piglets. The incidence of diarrhea post-challenge in the antibiotic and cecropin AD treatments decreased compared with the control piglets. The total viable counts of cecal E. coli were lower while the Lactobacilli counts were higher in the antibiotic and cecropin AD treatments compared with the control (P<0.05). Cecropin AD treatment decreased total aerobes while increasing total anaerobes in the ileum (P<0.05). A higher villus height to crypt depth ratio in the jejunum and ileum as well as a deeper crypt depth in the jejunum and higher villus height in the ileum were observed in piglets fed antibiotics or cecropin AD compared with control piglets (P<0.05). Piglets fed the control diet had lower levels of secretory IgA in their jejunum and lower serum IgA, IgG, interleukin-1ß and interleukin-6 compared with the other treatments (P<0.05). Overall, these data suggest that cecropin AD enhances pig performance through increasing immune status and nitrogen and energy retention as well as reducing intestinal pathogens in weaned piglets.