sw872 htb 92  (ATCC)

Journal: Clinical and Translational Medicine

Article Title: Oncogenic driver and therapeutic target: Prolactin signalling axis in retroperitoneal sarcoma

doi: 10.1002/ctm2.70669

Figure Lengend Snippet: Systematic identification of PRL‐regulating transcriptional machinery in liposarcoma. (A and B) Intersection analysis of candidate transcription factors (TFs): (A) Venn diagram integrating TFs from mesenchymal stem cell (MSC) versus adipocyte differential genes, transcriptors in distal transcriptional regions of PRL gene, and transcriptors in proximal transcriptional regions of the PRL gene ( p > .05, fold change > 2). (B) Comparative intersection of TFs from PRL‐high versus PRL‐low (cutoff criteria: median) sarcomas, transcriptors in distal transcriptional regions of PRL gene and transcriptors in proximal transcriptional regions of the PRL gene. (C–E) Correlation analysis between PRL mRNA and (C) SOX4 ( n = 12), (D) SOX9 ( n = 12), (E) SNAI1 ( n = 12) in clinical specimens, Log FC = Log10 fold change (sample X to sample minimum), fold change sample X to sample minimum = power (2, −[Ct(X) − Ct(gapdh)] − [Ct(X) − Ct(gapdh)]max). (F) Pharmacological induction assay: PRL mRNA fold‐change in SW872 cells treated with adipogenic cocktails (IBMX.5 mM, insulin 5 µg/mL, rosiglitazone 2 µM, dexamethasone 1 µM) versus DMSO control ( n = 4). (G) qPCR analysis of PRL and candidate TFs under IBMX (.5 mM) treatment ( n = 4). (H) Tissue microarray validation: SOX4 protein expression quantification by immunohistochemical analysis in adipose tissue ( n = 30), RWDLPS ( n = 20) and RDDLPS ( n = 50). Data expressed as mean ± SD unless specified; ** p < .01, *** p < .001 by two‐tailed Student's t ‐test; ns: not significant.

Article Snippet: The SW872 (HTB‐92) and HT1080 (CCL‐121) cell lines were purchased from ATCC.

Techniques: Control, Microarray, Biomarker Discovery, Expressing, Immunohistochemical staining, Two Tailed Test

Journal: Clinical and Translational Medicine

Article Title: Oncogenic driver and therapeutic target: Prolactin signalling axis in retroperitoneal sarcoma

doi: 10.1002/ctm2.70669

Figure Lengend Snippet: Mechanistic elucidation of SOX4‐mediated transcriptional activation of PRL. (A) Genomic architecture of PRL regulatory elements, highlighting the distal exon 1a of PRL and the conserved SOX4 binding sequence AACAAAG. (B) Adipocyte‐specific SOX4 knockout efficiency validation in Adipo‐Cre; Sox4 knockout mice versus wild‐type littermates ( n = 4). (C and D) Concomitant reduction of (C) PRL protein (ELISA) and (D) mRNA (qPCR) in SOX4‐deficient adipose tissue ( n = 4). (E and F) ChIP analysis demonstrating SOX4 protein occupancy at −493 bp upstream of the PRL promoter in beige adipocytes (E) and 3T3‐L1 cells (F), n = 4. (G) Examination of the effects of SOX4 knockout and overexpression on SW872 cells. (H) Changes in PRL mRNA levels in SW872 cells after SOX4 knockout and overexpression, n = 3. (I) ChIP analysis revealing SOX4 protein occupancy at −6362 bp upstream of the PRL promoter in SW872 cells, n = 4. (J) Relative transcriptional activity of the PRL promoter and SOX4 binding site mutant promoters in 293T cells transfected with vector, SOX4, promoter–luc or Mut–promoter–luc constructs, n = 4. (K) Relative transcriptional activity of the PRL promoter and SOX4 binding site mutant promoters in SW872 cells transfected with vector or SOX4 constructs, n = 3. (L and M) Analysis of the transcriptional regulation of PRL by SOX4 in response to cAMP, and its analogs bucladesine and 8‐Bromo‐cAmp, n = 3. (N and O) qPCR was employed to assess the mRNA expression levels of PRL and SOX4 in bromocriptine‐treated SW872 and 94T778 cells, n = 3. (P) The relative concentration of PRL protein in the culture supernatant of bromocriptine‐treated SW872 cells was measured, n = 3. Data expressed as mean ± SD unless specified; * p < .05, ** p < .01, *** p < .001 by two‐tailed Student's t ‐test; ns: not significant.

Article Snippet: The SW872 (HTB‐92) and HT1080 (CCL‐121) cell lines were purchased from ATCC.

Techniques: Activation Assay, Binding Assay, Sequencing, Knock-Out, Biomarker Discovery, Enzyme-linked Immunosorbent Assay, Over Expression, Activity Assay, Mutagenesis, Transfection, Plasmid Preparation, Construct, Expressing, Concentration Assay, Two Tailed Test

Journal: Clinical and Translational Medicine

Article Title: Oncogenic driver and therapeutic target: Prolactin signalling axis in retroperitoneal sarcoma

doi: 10.1002/ctm2.70669

Figure Lengend Snippet: Functional characterisation of PRL‐mediated proliferation and chemoresistance in sarcoma models. (A and B) Secretory PRL quantification by ELISA confirming knockdown efficiency in the culture medium, n = 3. (C–F) Growth suppression following PRL depletion: CCK‐8 time‐course assay ( n = 5) and colony formation capacity ( n = 3) in PRL‐knockdown models. (G–J) Recombinant PRL (50 ng/mL)‐induced proliferative enhancement: (G and H) CCK‐8 ( n = 5) and (I and J) colony formation ( n = 3) in HT1080 and SW872 lines. (K–P) PRLR‐dependent proliferation modulation: (K–N) CCK‐8 dose‐response ( n = 5) and (O‐P) colony formation ( n = 3) analysis post‐PRLR perturbation. (Q and R) After treating SW872 and HT1080 cells with PRLR antibody rolinsatamab talirine (20 µg/mL), the effect on cell proliferation was detected by the CCK8 method, with n = 5. (S and T) Xenograft tumourigenesis assay demonstrating impaired SW872 growth with PRL knockdown ( n = 9). (U) A single SW872 clone exhibiting the lowest PRL expression among the pooled PRL‐knockout cells was isolated by limiting dilution cloning, expanded in culture and validated for PRL protein levels via ELISA. (V) Cell proliferation was assessed using the CCK‐8 assay following stable PRL knockout ( n = 5 biological replicates). (W) Bromocriptine‐mediated antiproliferative effects were evaluated in parallel in wild‐type and PRL‐knockout SW872 cell lines using the CCK‐8 assay ( n = 5). (X) In vivo efficacy was determined in a subcutaneous xenograft mouse model, wherein tumour growth derived from wild‐type or PRL‐knockout SW872 cells was monitored following bromocriptine treatment, n = 7. (Y and Z) Chemosensitisation effects: PRL pretreatment (50 ng/mL) enhances cytotoxicity of RG7112/abemaciclib/doxorubicin/gemcitabine, n = 5, RG7112 (10 µM), abemaciclib (5 µM), doxorubicin (2 µM), gemcitabine (10 µM). (a) Western blot analysis was performed to detect MDM2 expression in human adipocytes, liposarcoma cell lines (SW872, 93T449, 94T778), fibrosarcoma cell line HT1080 and clinically isolated liposarcoma cell lines established in our laboratory. (b) Western blot analysis was performed to detect MDM2 in 12 clinical retroperitoneal liposarcoma tissues and its corresponding paracancerous tissues, 6 clinical retroperitoneal fibrosarcoma tissues and corresponding paracancerous tissues. (c) Therapeutic synergy evaluation: bromocriptine combined with RG7112 in WEHI164 fibrosarcoma murine model, RG7112: 100 mg/kg per day, bromocriptine: 10 mg/kg, twice daily, ( n = 6). Data expressed as mean ± SD unless specified; * p < .05, ** p < .01, *** p < .001 by two‐tailed Student's t ‐test; ns: not significant.

Article Snippet: The SW872 (HTB‐92) and HT1080 (CCL‐121) cell lines were purchased from ATCC.

Techniques: Functional Assay, Enzyme-linked Immunosorbent Assay, Knockdown, CCK-8 Assay, Recombinant, Expressing, Knock-Out, Isolation, Cloning, In Vivo, Derivative Assay, Western Blot, Two Tailed Test

Journal: Clinical and Translational Medicine

Article Title: Oncogenic driver and therapeutic target: Prolactin signalling axis in retroperitoneal sarcoma

doi: 10.1002/ctm2.70669

Figure Lengend Snippet: Analysis of the PRL‐regulated pathway. (A) OPLS‐DA score plot distinguishing PRL‐activated (12 h) versus control transcriptional profiles, C: control, P: prolactin, n = 4. (B) Volcano plot analysis. Red denotes up‐regulated genes, blue denotes down‐regulated genes and grey denotes insignificant changed genes. (C) Heatmap: a red colour indicates a higher expression level, while blue colour indicates a lower expression level. (D) KEGG pathway enrichment. (E) Regulatory network mapping PRL‐associated genes (circles) to tumour‐related pathways (squares). (F–L) Tissue microarray validation: IHC staining was performed on RLPS tissue and adipose tissue microarray using c‐MYC antibodies. Positive cell%, histochemistry SCORE and IRS are calculated. Adipose tissue: n = 30; retroperitoneum well‐differentiated liposarcoma (RWDLPS): n = 20; retroperitoneum dedifferentiated liposarcoma (RDDLPS): n = 50. T: sarcoma tissue, N: adipose tissue. (M and N) Clinical correlation: pre‐operative serum up‐regulates c‐MYC versus post‐operative serum in SW872. (O) Rescue experiment: PRL‐induced c‐MYC up‐regulation blocked by PRLR knockdown. (P) Western blot analysis of key proteins in the JAK–STAT signalling pathway were detected following down‐regulation of PRLR and addition of recombinant PRL (50 ng/mL) in SW872 cells. (Q) qPCR was performed to assess mRNA expression levels of PRL, STAT5A, STAT5B and MYC in SW872 cells following treatment with the cAMP agonist bucladesine or the STAT5 inhibitor pimozide, alone or in combination. Bucladesine treatment reversed pimozide‐induced suppression of MYC expression, Buc: bucladesine, Pi: pimozide, n = 3. (R) Combined treatment: PRL reverses c‐MYC inhibitor (APTO‐253, 10058‐F4) mediated c‐MYC induction. (S) Therapeutic synergy: cell survival rates were measured after individual or combined treatment with the MDM2 inhibitor RG7112, PRL recombinant protein and c‐MYC inhibitors (APTO‐253, 10058‐F4), n = 5. Data expressed as mean ± SD unless specified; ** p < .01, *** p < .001 by two‐tailed Student's t ‐test; ns: not significant.

Article Snippet: The SW872 (HTB‐92) and HT1080 (CCL‐121) cell lines were purchased from ATCC.

Techniques: Control, Expressing, Microarray, Biomarker Discovery, Immunohistochemistry, Knockdown, Western Blot, Recombinant, Two Tailed Test

Journal: Adipocyte

Article Title: mTORC2 regulates lipid metabolism-driven TAMs via the PPAR-γ/CD36 pathway to promote liposarcoma progression

doi: 10.1080/21623945.2026.2665903

Figure Lengend Snippet: IL-4-Induced M2 macrophage polarization promotes the progression of LPS. RAW264.7 cells were divided into two groups: control group, IL-4 group. (A) Flow cytometry was used to detect the expression level of CD206 on RAW264.7 cells in each group. (B) ELISA was performed to measure the secretion level of IL-10 in RAW264.7 cells from each group. (C) Western Blot was employed to determine the protein expression level of Arg-1 in RAW264.7 cells of each group. A Transwell co-culture system was utilized, which included two groups: control co-culture group (LPS cells co-cultured with untreated RAW264.7 cells). IL-4 co-culture group (LPS cells co-cultured with RAW264.7 cells pre-treated with IL-4). (D) EdU incorporation assay was used to detect the proliferative activity of SW872 cells and 94T778 cells in each group. (E-F) Transwell migration and invasion assays were performed to evaluate the migration and invasion abilities of SW872 cells and 94T778 cells in each group. Data are presented as the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. Experiments were repeated independently at least three times.

Article Snippet: Human liposarcoma cell lines SW872 and 94T778, as well as the murine macrophage cell line RAW264.7, were purchased from the American Type Culture Collection (ATCC, USA).

Techniques: Control, Flow Cytometry, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot, Co-Culture Assay, Cell Culture, Activity Assay, Migration

Journal: Adipocyte

Article Title: mTORC2 regulates lipid metabolism-driven TAMs via the PPAR-γ/CD36 pathway to promote liposarcoma progression

doi: 10.1080/21623945.2026.2665903

Figure Lengend Snippet: mTORC2 regulates TAM via PPAR-γ/CD36 pathway to promote LPS progression. LPS cells (SW872 and 94T778) were co-cultured with RAW264.7 macrophages, and the groups were divided as follows: control co-culture group, IL-4 co-culture group, IL-4+JR-AB2-011 co-culture group, IL-4+JR-AB2-011+LPA co-culture group. (A) EdU assay was used to detect the proliferation of SW872 and 94T778 cells in each co-culture group. (B-C) Transwell migration and invasion assays were performed to evaluate the migration and invasion abilities of SW872 and 94T778 cells in each co-culture group. Data are presented as the mean±SD. * p < 0.05, ** p < 0.01, *** p < 0.001. Experiments were repeated independently at least three times.

Article Snippet: Human liposarcoma cell lines SW872 and 94T778, as well as the murine macrophage cell line RAW264.7, were purchased from the American Type Culture Collection (ATCC, USA).

Techniques: Cell Culture, Control, Co-Culture Assay, EdU Assay, Migration

Journal: Cancer Medicine

Article Title: Clinical Significance and Therapeutic Potential of Long Non‐Coding RNA H19 in Soft Tissue Sarcoma

doi: 10.1002/cam4.71305

Figure Lengend Snippet: H19 expression in different sarcoma subtypes. (A) H19 RNA‐seq expression data across cell lines from 38 different cancer types derived from the publicly available CCLE database. (B) H19 RNA‐seq expression data from 31 cancer types in tumor tissue derived from TCGA and GTEx data (SARC = sarcoma; TMP = Transcript Per Million). (C) The expression of H19 in 7 different sarcoma cell lines was measured by qRT‐PCR and normalized to the housekeeper genes GAPDH and U6 ( n = 3; mean ± SD). (D) Representative pictures of RNA in situ hybridization of H19 in the liposarcoma cell line SW872 showing a heterogenous expression pattern.

Article Snippet: The sarcoma cell lines SW872 (Liposarcoma) and SW982 (Synovial Sarcoma) were purchased from the American Type Culture Collection (ATCC; Manassas, CA, USA).

Techniques: Expressing, RNA Sequencing, Derivative Assay, Quantitative RT-PCR, RNA In Situ Hybridization