Journal: Nature

Article Title: ACLY inhibition promotes tumour immunity and suppresses liver cancer

doi: 10.1038/s41586-025-09297-0

Figure Lengend Snippet: a , Volcano plot of bulk RNA-seq of tumours showing upregulated and downregulated genes at early or late timepoints from Acly- KO ( n = 12) versus WT ( n = 9) mice. Significance was determined by Wald test with a false discovery-adjusted threshold of 5% as implemented in DESeq2. Horizontal dashed lines demarcate the P value threshold at a 5% false discovery rate (FDR). b , Gene Ontology analysis of selected biological processes involving significantly upregulated genes in tumours from Acly -KO ( n = 12) versus WT ( n = 9) mice. FC, fold change. c , Top 10 Gene Ontology biological processes (GOBP) from clusters identified among significantly upregulated gene sets in tumours from Acly -KO ( n = 12) versus WT ( n = 9) mice. NES, normalized enrichment score. d , Correlation between B cell populations and Acly expression in tumours from WT ( n = 9) and Acly -KO ( n = 12) mice. Confidence bands denote the upper and lower bounds of the 95% confidence interval. Significance of association was determined by a two-sided Student’s t -test of regression coefficients and at a false discovery-adjusted threshold of 5%. e – p , Spatial transcriptomic analysis of livers from WT and Acly -KO mice and vehicle or EVT0185-treated mice. Cluster analysis representing the number of cell types in the liver and tumour ( e , k ). Umapharmony integration analysis showing increased B cells in Acly- KO ( f ) and in EVT0185-treated ( l ) mice. The top upregulated pathways in HCC cells from Acly -KO ( g ) and EVT0185-treated ( m ) mice. Statistical analysis was performed using Fisher’s exact test. Expression level of metabolic genes ( h , n ). Expression of markers of subtypes of B cells ( i , o ). The box-and-whisker plots are defined by the median with the first quartile (Q1), third quartile (Q3), minimum (Q1 − 1.5 × interquartile range (IQR)) and maximum (Q3 + 1.5 × IQR). Cxcl13 expression levels in HCC cells ( j , p ). GC, germinal centre; HSC, hepatic stellate cell; MZ, marginal zone; Tr, regulatory; VSMC, vascular smooth muscle cell.

Article Snippet: Liver tumour CXCL13 protein levels were determined using the mouse CXCL13/BLC/BCA-1 Quantikine ELISA Kit (MCX130, R&D Systems) following the manufacturer’s instructions.

Techniques: RNA Sequencing, Expressing, Whisker Assay

Journal: Nature

Article Title: ACLY inhibition promotes tumour immunity and suppresses liver cancer

doi: 10.1038/s41586-025-09297-0

Figure Lengend Snippet: a-b , Immune cell markers expressed in different cell types in a , WT and Acly KO and b , Vehicle and EVT0185-treated mice. c , Top upregulated pathways in B cells in Acly KO or EVT0185-treated mice (spatial transcriptomics analysis). Statistical analysis was performed using Fisher’s Exact test. d and e , Single seq analysis of d , WD-DEN and e , WD-CCl 4 mouse livers showing top upregulated pathways in B cells. Statistical analysis was performed using a one-sided hypergeometric test (enrichGO); p-values adjusted by Benjamini-Hochberg. f , Cxcl13 mRNA expression analyzed from publicly available RNA-seq dataset in WT and Acly KO DEN tumors cultured in vitro ( GSE223966 ) . Boxplot lines represent the first quartile, median, and third quartile. Whiskers connect the minimum and maximum values. Significance was ascertained by an unpaired two-tailed t-test between Acly KO vs WT (n = 4 hepatocellular carcinoma cell lines derived from DEN-induced tumors in Acly f/f mice).

Article Snippet: Liver tumour CXCL13 protein levels were determined using the mouse CXCL13/BLC/BCA-1 Quantikine ELISA Kit (MCX130, R&D Systems) following the manufacturer’s instructions.

Techniques: Expressing, RNA Sequencing, Cell Culture, In Vitro, Two Tailed Test, Derivative Assay

Journal: Nature

Article Title: ACLY inhibition promotes tumour immunity and suppresses liver cancer

doi: 10.1038/s41586-025-09297-0

Figure Lengend Snippet: a and b , Quantification of lipid droplet area was extrapolated by converting the PLIN-2 signal into a mask and quantifying the area covered by this mask per 400 × 400 micron field of view (FOV). These calculations were performed in ImageJ using custom scripts. Each dot represents a single FOV, Wilcoxon non-parametric unpaired two-sided test. Acly KO (n = 188) vs WT (n = 170) FOVs from n = 4 mice/group. c , CXCL13 protein level in tumors from WT and Acly KO mice; mean ± SEM, n = 5 mice/group; unpaired two-tailed t-test. d , Representative images showing B cells at tumor centre and periphery (Orange star: Tumor front (border-periphery), Black star: Surrounding non-tumoral hepatic tissue). e , B cell aggregations in Acly KO tumor (Orange star: Tumor, Black star: Non-tumoral liver tissue, Blue star: B Cell aggregations). f , Heatmap of cell neighborhoods hierarchically sorted by cell phenotype and showing relative cell abundances. ( g-j, l-n ) WD-DEN mice treated with Vehicle or EVT0185. g , Representative MIBI images of liver tumors. h , B cell, and i , T cell counts from regions representing the tumor lesion interface with the liver; mean ± SEM, n = 4 mice/group; unpaired Wilcoxon non-parametric unpaired two-sided test. j , CD19+ positive cells count/mm 2 tumor area; mean ± SEM, Vehicle (n = 29) and EVT0185 (n = 21) lesions; unpaired two-tailed t-test. k , CD3+ positive cells count/mm 2 tumor area; mean ± SEM, WT (n = 12) and Acly KO (n = 10) lesions; unpaired two-tailed t-test. l , CD3+ positive cells count/mm 2 tumor area; mean ± SEM, Vehicle (n = 29) and EVT0185 (n = 21) lesions; unpaired two-tailed t-test. m , Percentage of mice with/without B cells in TLS. n , TLS in EVT-treated mouse compared with vehicle control. o , GO enrichment analysis showing upregulated pathways related to complement activation in B cells in the EVT0185-treated group relative to Vehicle. Statistical analysis was performed using one-sided hypergeometric test (enrichGO); p-values adjusted by Benjamini-Hochberg.

Article Snippet: Liver tumour CXCL13 protein levels were determined using the mouse CXCL13/BLC/BCA-1 Quantikine ELISA Kit (MCX130, R&D Systems) following the manufacturer’s instructions.

Techniques: Two Tailed Test, Control, Activation Assay

Journal: Nature

Article Title: ACLY inhibition promotes tumour immunity and suppresses liver cancer

doi: 10.1038/s41586-025-09297-0

Figure Lengend Snippet: a , Differential expression analysis comparing MASH-HCC (n = 53) and non-tumor adjacent liver (n = 29), MASH liver (n = 74), cirrhotic liver (n = 8), and healthy liver tissue (n = 6); two-tailed t-test; 5% FDR as implemented in limma. b , Overlap between significantly upregulated and downregulated genes in each pairwise comparison. c , Gene expression of lipogenic enzymes acetyl-CoA carboxylase ( ACACA or ACACB ), ACSS2 , and FASN across disease states (MASH-HCC, n = 53; non-tumor adjacent, n = 29; MASH liver, n = 74; cirrhotic liver, n = 8; healthy liver, n = 6). Boxplot lines represent the first quartile, median, and third quartile. Whiskers connect the minimum and maximum values. d , ACLY upregulation in human MASH-HCC compared to all other tissue types using scRNAseq samples. Significance assessed using Wilcoxon rank-sum tests comparing each condition to MASH-HCC, with p-values adjusted using the Benjamini-Hochberg method. Exact p-adj values: vs healthy P adj = 6.9 × 10 −247 , vs MALSD P adj = 1.0 × 10 −16 , vs non-tumor adjacent P adj = 1.6 × 10 −18 . e , Differential expression analysis of genes associated with ACLY expression in MASH-HCC; two-tailed t-test; 5% FDR as implemented in limma. f , Principal component embeddings of genes differentially expressed with respect to ACLY expression validate stratification of MASH-HCC tissues by high, medium, and low ACLY expression levels. g , Upregulation of CXCL13 among human MASH-HCC samples with reduced (bottom tertile, n = 18) relative to elevated (top tertile, n = 18) ACLY expression. Significance determined by two-tailed t-test with a false discovery adjusted threshold of 5% as implemented in limma. Boxplot lines represent the first quartile, median, and third quartile. Whiskers connect the minimum and maximum values. h , Identification of co-expression modules associated with ACLY expression and immune features. i , Significant biological processes associated with the immune and ACLY associated gene co-expression module.

Article Snippet: Liver tumour CXCL13 protein levels were determined using the mouse CXCL13/BLC/BCA-1 Quantikine ELISA Kit (MCX130, R&D Systems) following the manufacturer’s instructions.

Techniques: Quantitative Proteomics, Two Tailed Test, Comparison, Gene Expression, Expressing

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: (A) Protein–protein intersection (PPI) network of differentially expressed genes (DEGs); (B) twenty core genes in the PPI network; (C) prognostic DEGs; (D) core genes CXCL13 and CD3E on the Venn diagram; (E) meta-analysis for OS based on expression levels of CXCL13 in OC patients.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: TCGA and GETx datasets show expression of CXCL13 is statistical different between OC and normal tissue (A) and in different pathological stages of OC (B). (C) Oncomine database verifies the expression of CXCL13 ; (D) the negative staining of CXCL13 in 2 normal tissues; (E) the negative (0), low (1), and medium (2) immunohistochemical staining of CXCL13 in 3 OC patients.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing, Negative Staining, Immunohistochemical staining, Staining

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: Survival curves of OC patients in CXCL13 high- and low-expression groups.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: Gene set enrichment analysis enrichment pathway of CXCL13 expression profile in OC patients.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: Correlation between the expression of CXCL13 and infiltration of various immune cell components in the tumor microenvironment. (A) CD8 + T cell; (B) activated CD4 + memory T cell; (C) memory B cell; (D) M1 macrophage cell; (E) follicular helper T cell; (F) resting CD4 + memory T cell; (G) naive B cell; (H) M2 macrophage cell; (I) activated myeloid dendritic cell; (J) neutrophils.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing

Journal: Medicine

Article Title: Study on correlation between CXCL13 and prognosis and immune characteristics of ovarian cancer

doi: 10.1097/MD.0000000000040272

Figure Lengend Snippet: (A) Correlation between the expression of CXCL13 and PD-1 of TCGA-OC samples; (B) correlation between the expression of CXCL13 and CR/PR, SD/PD in mUC samples after receiving PD-L1 blocking therapy (CR = complete response, PR = partial response, PD = progressive disease, SD = stable disease). Survival curves of mUC patients with high- and low- CXCL13 expression in SD/PD (C) and CR/PR (D) group.

Article Snippet: The Human Protein Atlas (HPA) ( http://proteinatlas.org ) [ ] was used to analyze CXCL13 protein levels.

Techniques: Expressing, Blocking Assay

Journal: Frontiers in Immunology

Article Title: Role of CXCL5 in Regulating Chemotaxis of Innate and Adaptive Leukocytes in Infected Lungs Upon Pulmonary Influenza Infection

doi: 10.3389/fimmu.2021.785457

Figure Lengend Snippet: CXCL5 regulated CXCL13 expression in pulmonary CD64+ macrophages/monocytes via the CXCL1/2/5-CXCR2 signaling pathway. (A–C) The protein levels of CXCL1/2 (A) , CXCL12 (B) , and CXCL13 (C) in the BALF of infected mice were measured by ELISA, and the level of CXCL5 was used as a reference (n=5). (D) RT-qPCR for IFN-α and IFN-β mRNA was performed in lung homogenates from WT and CXCL5 -/- mice before and after (3 d.p.i.) influenza infection. The fold changes in the mRNA expression levels of these genes were calculated using the 2 -ΔΔCt method of relative quantification with GAPDH as the endogenous reference gene. The relative fold changes in IFN-α and IFN-β expression in infected mice compared to normal mice are presented (n=4). (E, F) Percentages and numbers of CXCL13-expressing cells among single lung cells from WT and CXCL5 -/- mice at 3 d.p.i. (E) and 8 d.p.i. (F) as assessed by intracellular staining of CXCL13 protein (APC-CXCL13) and flow cytometry (n=4). (G, H) Percentages and numbers of CXCL13-expressing cells among pulmonary CD64+ cells from WT and CXCL5 -/- mice at 3 d.p.i. (G) and 8 d.p.i. (H) as assessed by intracellular staining of CXCL13 protein (APC-CXCL13) and flow cytometry (n=4). (I) Percentages and numbers of CXCL13-expressing cells among pulmonary CD11c+ cells from WT and CXCL5 -/- mice at 8 d.p.i. as assessed by intracellular staining of CXCL13 protein (APC-CXCL13) and flow cytometry (n=4). (J–L) Percentages and numbers of CD206 (J) , CD44 (K) , and CD274 (L) surface markers among pulmonary CD64+ cells from WT and CXCL5-/- mice at 3 d.p.i. (n=4). (M) Detection of the expression of mouse CXCR2 on total AMs and CD64+ AMs by western blotting. (N) RT-qPCR was performed on cell lysates of cultured CD64+ macrophages after 10 h of influenza challenge or unstimulated (control) conditions, and the fold changes were calculated as described in the MATERIALS AND METHODS section (n=4). (O) Expression levels of the indicated chemokines from supernatants of cultured CD64+ macrophages after 20 h of influenza challenge or unstimulated (control) conditions as determined by ELISA (n=4). (P) ELISA was used to determine the expression levels of CXCL13 protein in supernatants of cultured CD64+ macrophages after 20 h of influenza challenge and treatment with the indicated recombinant chemokine proteins, the CXCR2 antagonist SB225002, the PI3K inhibitor LY294002, and the MEK inhibitor PD98059 (n=3). The error bars represent the SDs. * P < 0.05 based on Student’s t -test.

Article Snippet: CXCL5 (MX000), CXCL1 (MKC00B), CXCL2 (MM200), CXCL12 (MCX120), and CXCL13 (MCX130) levels were determined using ELISA kits from R&D Systems (MN, USA) in accordance with the manufacturer’s instructions.

Techniques: Expressing, Infection, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Quantitative Proteomics, Staining, Flow Cytometry, Western Blot, Cell Culture, Control, Recombinant

Journal: Frontiers in Immunology

Article Title: Role of CXCL5 in Regulating Chemotaxis of Innate and Adaptive Leukocytes in Infected Lungs Upon Pulmonary Influenza Infection

doi: 10.3389/fimmu.2021.785457

Figure Lengend Snippet: CXCL5-CXCR2 signaling regulated pulmonary neutrophil and B cell recruitment and CXCL13 production from lung macrophages upon H1N1 infection. (A–E) WT mice were infected with H1N1 virus (1000 CCID 50 ) and treated with either CXCR2 antagonist or PBS as a control during 1-3 d.p.i. Microscopic view of BALF leukocytes from the infected mice collected at 3 d.p.i. and cytospun (×400 magnification) (A) . The protein level of CXCL13 in the BALF of the infected mice was measured at 3 d.p.i. by ELISA (n=4) (B) . Percentages and numbers of CXCL13-expressing cells among pulmonary CD64+ cells (C) , CD19+ B cells (D) , and CD44+/CD274+ CD64+ cells (E) from the infected mice at 3 d.p.i. by flow cytometry and counting (n=4). (F–J) WT mice were infected with H1N1 virus (1000 CCID 50 ) and orotracheally instilled with either CXCL5 recombinant protein or PBS as a control during 5-8 d.p.i. Microscopic view of BALF leukocytes from the infected mice collected at 8 d.p.i. and cytospun (×400 magnification) (F) . The protein level of CXCL13 in the BALF of the infected mice was measured at 8 d.p.i. by ELISA (n=4) (G) . Percentages and numbers of CD19+ B cells from the infected mice at 8 d.p.i. by flow cytometry and counting (n=4) (H) . Measurements of H1N1-specific IgG (I) and IgA (J) in the BALF of the CXCL5 treatment and control mice at 8 d.p.i. (n = 4). The error bars represent the SDs of 4 samples. * P < 0.05 based on Student’s t -test.

Article Snippet: CXCL5 (MX000), CXCL1 (MKC00B), CXCL2 (MM200), CXCL12 (MCX120), and CXCL13 (MCX130) levels were determined using ELISA kits from R&D Systems (MN, USA) in accordance with the manufacturer’s instructions.

Techniques: Infection, Virus, Control, Enzyme-linked Immunosorbent Assay, Expressing, Flow Cytometry, Recombinant