protein profile determination  (Bio-Rad)

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A Experimental design of an in vivo high-throughput screening for anti-tumor immunotherapeutic agents using Drosophila stem cell-derived tumor model. Created in BioRender. Lee, H. (2026) https://BioRender.com/4zb6pi7 . B Representative images showing the immunofluorescent staining of GFP + Arm + Ras V12 -transformed renal nephric stem cells in Drosophila treated with DMSO or STK. C Quantification of the tumor size of renal nephric stem cell tumors in ( B ) ( n = 12 biologically independent samples). D Experimental design of strategies to identify target proteins of STK. The reverse virtual screening was performed by using Pharmmapper and Chemmapper databases for pharmacophore matching and potential target identification, followed by Drug Affinity Responsive Target Stability (DARTS) experiments. Created in BioRender. Lee, H. (2026) https://BioRender.com/lb2oxp7 . E An Venn diagram of potential target proteins identified by virtual screening and DARTS. F Immunoblotting analysis of DARTS detecting the binding of STK to SOAT1. Data are representative of three independent experiments ( n = 3 biological replicates). G Molecular docking analysis of STK binding to human SOAT1. H Isothermal titration calorimetry (ITC) analysis of binding of STK to SOAT1 purified from E. coli . I , J Tumor growth curves and tumor weight of BALB/c mice transplanted with Scram-knockdown or Soat1 -knockdown CT26 cells ( n = 5 biologically independent samples). Data are shown as the mean ± SD. Scale bars are as indicated. Statistical significance was assessed by a two-tailed unpaired Student’s t -test or two-way ANOVA. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: In Vivo, High Throughput Screening Assay, Derivative Assay, Staining, Transformation Assay, Drug discovery, Western Blot, Binding Assay, Isothermal Titration Calorimetry, Purification, Knockdown, Two Tailed Test

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A Schematic diagram of treatment design for Lgr5-CreER-GFP/Apc f/f mice. Created in BioRender. Lee, H. (2026) https://BioRender.com/ol3f47g . B Representative images of immunohistochemistry staining of cancer stem cells in the intestine of CSC-like colon cancer mice treated with DMSO or STK ( n = 12 mice in each group). C Quantification of tumor numbers in ( B ) ( n = 12 biologically independent samples). D Schematic diagram of treatment design for MYC-ON mice. Created in BioRender. Lee, H. (2026) https://BioRender.com/0wx33vy . E Representative liver images of MYC-ON mice treated with DMSO (left) or STK (right). F Tumor numbers of MYC-ON mice treated with DMSO or STK. ( n = 8 biologically independent samples). G Survival curves of MYC-ON mice treated with DMSO or STK. ( n = 6 biologically independent samples). H Schematic diagram of treatment design for LSL-Kras/P53 f/f mice. Created in BioRender. Lee, H. (2026) https://BioRender.com/kh1x47x . I Representative lung images of lung adenocarcinoma mice treated with DMSO (left) or STK (right). J Tumor numbers of lung cancer mice treated with DMSO or STK. ( n = 10 mice in each group). K Survial curves of lung cancer mice treated with DMSO or STK. ( n =8 mice in each group). L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates). Images shown are representative results from multiple tissue samples. O – S Kaplan–Meier analysis of the correlation between SOAT1 expression levels and overall survival (OS) of patients from the TCGA dataset. T Immunoblotting analysis of SOAT1 levels in the stem CT26 subpopulation and non-stem CT26 subpopulation. U Flow cytometry analysis of SOAT1 expression levels between normal and tumor tissues from human lung carcinoma patients. V Flow cytometry analysis of SOAT1 expression levels on stem cancer cells or non-stem cancer cells from human lung carcinoma patients. W Immunofluorescence (IF) staining of tumor tissues from CT26 subcutaneous tumor-bearing mice ( n = 3 biological replicates). Orange: CD133; Green: SOAT1; Blue: DAPI. Yellow arrowheads indicate cells with co-localization of CD133 and SOAT1. Scale bar: 10 µm. Shown is a representative image from three independent experiments. Data are shown as the mean ± SD or the Kaplan-Meier test. Scale bars are as indicated. Statistical significance was assessed by a two-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: Immunohistochemistry, Staining, Expressing, Derivative Assay, Western Blot, Flow Cytometry, Immunofluorescence, Two Tailed Test

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A Volcano plot of differentially expressed genes in tumor-infiltrated Tregs from live cancer mice treated with or without STK. B NicheNet ligand-receptor analysis plot between tumor cells and Tregs from lung cancer mice treated with or without STK. C Representative flow cytometry plots of GPR132, CTLA-4 and LAG3 subpopulation in Tregs in LSL-Kras/P53 f/f /Soat1 f/f mice or LSL-Kras/P53 f/f mice. D – F Quantification of GPR132, CTLA-4 and LAG-3 positive percentage in LSL-Kras/P53 f/f /Soat1 f/f mice or LSL-Kras/P53 f/f mice. ( n = 3 in each group) ( G , H ) Kaplan–Meier analysis of the correlation between GPR132, FOXP3 and CTLA-4 expression levels and overall survival (OS)/disease-free survival (DFS) in patients from the TCGA dataset. I , J Tumor growth curves of BALB/c mice transplanted with CT26 cells with or without anti-CD25 antibody treatment, in combination with DMSO or STK. ( n = 5 biologically independent samples). K – N Tumor growth curves and tumor weight in Foxp3 Cre mice (K and L) or Foxp3 Cre ; Gpr132 f/f mice (M and N). LLC cells transfected with ShScram or Sh Soat1 were transplanted into Foxp3 Cre or Foxp3 Cre ; Gpr132 f/f mice ( n = 5 biologically independent samples). Data are shown as the mean ± SD. Statistical significance was assessed by a two-tailed unpaired Student’s t -test or two-way ANOVA. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: Flow Cytometry, Expressing, Transfection, Two Tailed Test

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A , B Heatmap of pathway enrichment analysis of tumor-infiltrated Tregs from liver cancer mice treated with or without STK, based on scRNA-seq data. C Proposed model of GPR132 signaling pathway in Tregs. Created in BioRender. Lee, H. (2026) https://BioRender.com/ou98v3q . D , F Secretion levels of TGF-β and IL-10 in Tregs cultured with conditional medium (CM) from DMSO-treated or STK-treated CT26 cells. ( n = 13 or 9 in each group). E , G Secretion levels of TGF-β and IL-10 in Tregs cultured with conditioned medium from the Scramble -knockdown or Soat1 -knockdown CT26 cells. ( n = 6 in each group). H , I Secretion levels of TGF-β in Tregs cultured with conditioned medium from indicated cells treated with GPR132 inhibitor telmisartan or STAT1 inhibitor fludarabine. ( n = 3 or 4 in each group). J , K Secretion levels of IL-10 in Tregs cultured with conditioned medium from indicated cells treated with GPR132 inhibitor telmisartan or STAT1 inhibitor fludarabine. ( n = 3 or 4 in each group). L – O Expression of GPR132 and LAG-3 on Tregs cultured in CM from STK or DMSO-treated CT26 cells. Tregs were concurrently treated with vehicle, GPR132 inhibitor telmisartan or STAT1 inhibitor fludarabine. Representative flow cytometry plots of GPR132(L) and LAG-3(N). Quantification of GPR132 (M) and LAG-3 (O) in Treg cells. ( n = 4 in each group). Data are shown as the mean ± SD. Statistical significance was assessed by a two-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: Cell Culture, Knockdown, Expressing, Flow Cytometry, Two Tailed Test

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A , B Flow cytometry analysis and statistical analysis of NBD-labeled cholesterol uptake. Tregs were cultured with conditioned medium (CM) collected from CT26 cells that had been pulsed with NBD-cholesterol and treated with DMSO or STK. ( n = 5 in each group). C Enrichment of gene sets related to oxysterol metabolism in tumor cells based on proteomic data. D Heatmap of oxysterol-related enzymes mRNA levels of bulk RNA sequencing from Hepa1-6 cells. ( n = 3 in each group). E Heatmap analysis of oxysterol-related enzymes mRNA levels in Hepa1-6 cells treated with DMSO or STK detected by qPCR. ( n = 3 in each group). F Heatmap analysis of mRNA of oxysterol-related enzymes in CT26 cells with Soat1 knockdown or treated with STK detected by qPCR. ( n = 3 in each group). G , H LC-MS analysis of oxysterols in the conditioned medium of Hepa1-6 or CT26 cells treated with DMSO or STK. I Immunoblotting analysis of DARTS-based detection for the binding of 20SOHC to GPR132. Data are representative of three independent experiments. J , K Molecular docking analysis of 20SOHC or 9S-HODE binding to human GPR132. Data are shown as the mean ± SD. Statistical significance was assessed by a two-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: Flow Cytometry, Labeling, Cell Culture, RNA Sequencing, Knockdown, Liquid Chromatography with Mass Spectroscopy, Western Blot, Binding Assay, Two Tailed Test

Journal: Nature Communications

Article Title: Inhibition of the cancer stem cell immune checkpoint SOAT1 suppresses regulatory T cell functions through a trans-cellular 20(S)-Hydroxycholesterol-GPR132 pathway in mice

doi: 10.1038/s41467-026-69305-3

Figure Lengend Snippet: A , D Representative tumor images of CT26 syngeneic tumor-bearing mice or 4T1 syngeneic tumor-bearing mice treated with DMSO or STK, in combination with anti-PD-1 blocking monoclonal antibodies (mAb). B , C Tumor growth curves and tumor weight of CT26 syngeneic tumor-bearing mice with indicated treatments ( n = 6 biologically independent samples). E , F Tumor growth curves and tumor weight of 4T1 syngeneic cancer mice with indicated treatments ( n = 5 biologically independent samples). G Experimental design of generating patient-derived xenograft (PDX) models of colon cancer in humanized immune system mice. Created in BioRender. Lee, H. (2026) https://BioRender.com/eu4e0mu . H , I Tumor growth curves and tumor weight of PDX mice with indicated treatment ( n = 6 biologically independent samples). J – M Flow cytometry analysis of the proportion of Tregs, the levels of GPR132 and CTLA4 in Tregs, and the proportion of CD8 + T cells in PDX mice ( n = 3 biologically independent samples). N Flow cytometry analysis of GPR132 expression in Tregs from paired lung carcinoma patient tumor samples and normal tissues. O The correlation analysis of SOAT1 expression in tumor cells and GPR132 expression levels in Tregs detected in lung carcinoma patient tumor samples. P , Q Kaplan–Meier analysis of the correlation between Cyp11a1 levels and overall survival (OS) in liver cancer patients from the TCGA dataset. Data are shown as the mean ± SD. Scale bars are as indicated. Statistical significance was assessed by a two-way ANOVA or two-tailed unpaired Student’s t -test. Source data are provided as a Source Data file.

Article Snippet: L – N SOAT1 protein expression profiles in paired normal and tumor tissues across human lung, colorectal and hepatic carcinomas derived from Human Protein Atlas (HPA) database ( n = 3 biological replicates).

Techniques: Blocking Assay, Bioprocessing, Derivative Assay, Flow Cytometry, Expressing, Two Tailed Test