a673 human ewing sarcoma cell line  (ATCC)

Journal: Molecular Therapy Oncology

Article Title: Trabectedin promotes oncolytic virus antitumor efficacy, viral gene expression, and immune effector function in models of bone sarcoma

doi: 10.1016/j.omton.2024.200886

Figure Lengend Snippet: oHSV+trabectedin synergizes to increase the disease control rate and reduce tumor burden in human xenograft models The best response for each treated tumor through 28 days, the average tumor burden, and spider plots tracking individual tumor volumes over the full study period are shown for (A) CHLA-258 (Ewing sarcoma), (B) EW5 (Ewing sarcoma), (C) PDX-0027 (rhabdomyosarcoma), (D) A673 (Ewing sarcoma), and (E) A673 in NSG-SGM3 NK-deficient mice (lack T, B, and NK cells). PBS and oHSV (1.0 × 10 7 pfu) were given intratumorally (i.Tu.) on days 0 and 2. Trabectedin (0.15 mg/kg) was given intravenously (i.v.) on days 0 and 7. Statistical analyses of the disease control rates (CR + PR + SD) were performed using a pairwise Fisher’s exact test with p values adjusted using the Benjamini-Hochberg procedure; ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001. Summarized data with error bars depict mean ± SEM.

Article Snippet: The A673 human Ewing sarcoma cell line (Cat# CCL-81), Vero green monkey kidney cell line (Cat# CRL-1598), and K7M2 mouse osteosarcoma cell line (Cat# CRL-2836) were purchased from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Control

Journal: Molecular Therapy Oncology

Article Title: Trabectedin promotes oncolytic virus antitumor efficacy, viral gene expression, and immune effector function in models of bone sarcoma

doi: 10.1016/j.omton.2024.200886

Figure Lengend Snippet: Trabectedin increases viral intracellular prevalence and decreases antiviral gene expression in immunodeficient Ewing sarcoma models (A) Treatment schema for scRNA-seq tumor collection: PBS and oHSV (1.0 × 10 7 pfu) were given i.Tu. on days 0 and 2, trabectedin (0.15 mg/kg) was given i.v. on day 0, and scRNA-seq samples were collected on day 3 to ensure sufficient cell viability for sequencing ( n = 2 per treatment group). (B) oHSV viral titer (pfu) from A673 tumors 3 days or 6 days following treatment with a single dose of oHSV or oHSV+trabectedin. Error bars indicate standard deviation and the p values from a Student’s unpaired t test between treatment groups at each time point are shown. (C) UMAP plots of oHSV transcript presence in all tumor cells, split by treatment group, from the merged scRNA-seq datasets. (D) Violin plot of the expression level (log-transformed) of all oHSV transcripts for all tumor cells in each treatment group. (E) Expression level of HSV time-dependent gene modules shown as a feature plot (UMAP) for all tumor cells and split by treatment groups that contained oHSV. A relative expression cutoff of 1 minimized color skewing by high-expressing outlier cells and maintained color scale consistency between feature plots. (F) Violin plot of the expression level (log-transformed) of HSV time-dependent gene modules for all tumor cells in treatment groups that contained oHSV. (G) Results from gene set enrichment analysis showing the running enrichment score for the “Herpes simplex virus 1 infection” pathway, which represents the intrinsic cellular response to HSV-1. Gene set enrichment analysis was performed on a gene list which was ranked by the gene expression fold change (log2FC) calculated for tumor cells treated with oHSV+trabectedin compared with those treated with oHSV monotherapy. The schema displays key genes from the “Herpes simplex virus 1 infection” pathway with their respective percent change shown in parentheses (a negative value indicates lower expression in tumors treated with oHSV+trabectedin, as compared to oHSV-treated tumors).

Article Snippet: The A673 human Ewing sarcoma cell line (Cat# CCL-81), Vero green monkey kidney cell line (Cat# CRL-1598), and K7M2 mouse osteosarcoma cell line (Cat# CRL-2836) were purchased from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Gene Expression, Sequencing, Standard Deviation, Expressing, Transformation Assay, Virus, Infection

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Broad spectrum activity of the checkpoint kinase 1 inhibitor prexasertib as a single agent or chemopotentiator across a range of preclinical pediatric tumor models

doi: 10.1158/1078-0432.CCR-18-2728

Figure Lengend Snippet: Relative EC50 values of prexasertib in a panel of pediatric cancer cell lines.

Article Snippet: Human pediatric sarcoma cell lines A673 (Ewing’s sarcoma, cat#CRL-1598), MG-63 (osteosarcoma, cat#CRL-1427), SJCRH30 (alveolar rhabdomyosarcoma [aRMS], cat# CRL-2061), and RD (embryonal RMS [eRMS], cat#CCL-136) were purchased from American Type Culture Collection (ATCC).

Techniques: Inhibition, Mutagenesis

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Broad spectrum activity of the checkpoint kinase 1 inhibitor prexasertib as a single agent or chemopotentiator across a range of preclinical pediatric tumor models

doi: 10.1158/1078-0432.CCR-18-2728

Figure Lengend Snippet: Animals bearing xenografts of pediatric bone or soft tissue sarcoma were treated with vehicle, prexasertib, chemotherapy, or the combination of prexasertib plus chemotherapy. The specific chemotherapy used is indicated in the key in each panel. A and B, SJCRH30 alveolar RMS CDX, n = 5/arm for each experiment. C, Rh41 alveolar RMS CDX, n = 4/arm. D, CCSARC005 osteosarcoma PDX, n = 5/arm, prexa* = dose reduced from 10 mg/kg to 8 mg/kg only in the combination due to use of SCID animals (this model only). E, A673 Ewing’s sarcoma CDX, n = 5/arm. For B and C, single experiments are displayed in two different graphs with the same vehicle and prexasertib arms but different chemotherapy and combination arms for clearer visualization. Dotted vertical lines: dosing interval; error bars: SEM. Waterfall plots for each model are displayed in Supplementary Fig. S4 and statistical analyses are summarized in Supplementary Table S3.

Article Snippet: Human pediatric sarcoma cell lines A673 (Ewing’s sarcoma, cat#CRL-1598), MG-63 (osteosarcoma, cat#CRL-1427), SJCRH30 (alveolar rhabdomyosarcoma [aRMS], cat# CRL-2061), and RD (embryonal RMS [eRMS], cat#CCL-136) were purchased from American Type Culture Collection (ATCC).

Techniques:

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Broad spectrum activity of the checkpoint kinase 1 inhibitor prexasertib as a single agent or chemopotentiator across a range of preclinical pediatric tumor models

doi: 10.1158/1078-0432.CCR-18-2728

Figure Lengend Snippet: Relative EC50 values of prexasertib in a panel of pediatric cancer cell lines.

Article Snippet: Cell culture conditions Human pediatric sarcoma cell lines A673 (Ewing’s sarcoma, cat#CRL-1598), MG-63 (osteosarcoma, cat#CRL-1427), SJCRH30 (alveolar rhabdomyosarcoma [aRMS], cat# CRL-2061), and RD (embryonal RMS [eRMS], cat#CCL-136) were purchased from American Type Culture Collection (ATCC).

Techniques: Inhibition, Mutagenesis

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Broad spectrum activity of the checkpoint kinase 1 inhibitor prexasertib as a single agent or chemopotentiator across a range of preclinical pediatric tumor models

doi: 10.1158/1078-0432.CCR-18-2728

Figure Lengend Snippet: Animals bearing xenografts of pediatric bone or soft tissue sarcoma were treated with vehicle, prexasertib, chemotherapy, or the combination of prexasertib plus chemotherapy. The specific chemotherapy used is indicated in the key in each panel. A and B, SJCRH30 alveolar RMS CDX, n = 5/arm for each experiment. C, Rh41 alveolar RMS CDX, n = 4/arm. D, CCSARC005 osteosarcoma PDX, n = 5/arm, prexa* = dose reduced from 10 mg/kg to 8 mg/kg only in the combination due to use of SCID animals (this model only). E, A673 Ewing’s sarcoma CDX, n = 5/arm. For B and C, single experiments are displayed in two different graphs with the same vehicle and prexasertib arms but different chemotherapy and combination arms for clearer visualization. Dotted vertical lines: dosing interval; error bars: SEM. Waterfall plots for each model are displayed in Supplementary Fig. S4 and statistical analyses are summarized in Supplementary Table S3.

Article Snippet: Cell culture conditions Human pediatric sarcoma cell lines A673 (Ewing’s sarcoma, cat#CRL-1598), MG-63 (osteosarcoma, cat#CRL-1427), SJCRH30 (alveolar rhabdomyosarcoma [aRMS], cat# CRL-2061), and RD (embryonal RMS [eRMS], cat#CCL-136) were purchased from American Type Culture Collection (ATCC).

Techniques:

Journal: Journal for Immunotherapy of Cancer

Article Title: Analysis of tumor-infiltrating NK and T cells highlights IL-15 stimulation and TIGIT blockade as a combination immunotherapy strategy for soft tissue sarcomas

doi: 10.1136/jitc-2020-001355

Figure Lengend Snippet: Combined IL-15 stimulation and TIGIT blockade augments killing of sarcomas in vitro when target expression of TIGIT ligands is high. Analysis of the TCGA-SARC database by Kaplan-Meier analysis shows a significant survival advantage in tumors with a (A) high NK signature (p=0.0003), (B) high CD8 signature (p=0.03), but independent of (C) TIGIT expression (p=0.17). (D) TIGIT expression within STS tumors has a strong positive correlation with NK signature (r=0.82, p<0.0001) and CD8 signature (r=0.90, p<0.0001). (E) Low expression of the primary TIGIT ligand CD155 was associated with improved overall survival by Kaplan-Meier analysis of the TCGA-SARC database (p=0.04). (F) Expression of TIGIT ligand CD155 on sarcoma cell lines A673 (red) and SK-LMS (blue) compared with FMO staining. (G) Increased cytotoxicity of CFSE-labeled target cells A673 (left) and SK-LMS (right) following 4-hour incubation with in vitro IL-15-preactivated PBMCs from a patient with sarcoma in the presence anti-TIGIT 10 ug/mL (red) compared with IgG control 10 ug/mL (blue) at low and high E:T ratios. Increased expression of degranulation marker CD107a on NK cells from IL-15-preactivated PBMCs following 4-hour incubation with sarcoma cell lines shown by (H) representative flow cytometry against A673 cells, and (I) quantified per cent expression against A673 and SK-LMS cell lines in a 5:1 E:T ratio. These experiments were repeated independently in two patients with STS with representative data shown. P value determined using Student’s t-test using n=3–4 technical replicates. FMO, Fluorescence Minus One; IL, interleukin; PBMCs, peripheral blood mononuclear cells; STS, soft tissue sarcoma; TCGA, The Cancer Genome Atlas.

Article Snippet: Human sarcoma cell lines A673 (#CRL-1598, Ewing’s sarcoma) and SK-LMS (#HTB-88, leiomyosarcoma) were obtained from ATCC (Manassas, Virginia, USA) and cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% Nu-Serum and 1% penicillin-streptomycin.

Techniques: In Vitro, Expressing, Staining, Labeling, Incubation, Control, Marker, Flow Cytometry, Fluorescence

Journal: American Journal of Cancer Research

Article Title: Acid microenvironment promotes cell survival of human bone sarcoma through the activation of cIAP proteins and NF-κB pathway

doi:

Figure Lengend Snippet: Pharmacological inhibition of the proton pump V-ATPase causes tumor necrosis in a Ewing’s sarcoma xenograft model. A. A673 cells were subcutaneously injected in NOD/SCID mice. Mice were then treated with different concentration of omeprazole, a V-ATPase inhibitor (0 mg/kg n = 14, 2.5 mg/kg n = 7, 10 mg/kg n = 14), to reduce intratumoral acidosis. H&E staining of tumor sections, representative images of the necrotic area, scale bar, 100 µm; B. Box plot of the tumor volume. Mean ± SE; C. Box plot of the tumor necrosis. Mean ± SE (*P < 0.05, **P < 0.01).

Article Snippet: HOS, Saos2, MG-63 (osteosarcoma), and A673 (Ewing sarcoma) human cell lines were purchased from the American Type Culture Collection, cultured in IMDM plus 20 unit/mL penicillin, 100 μg/mL streptomycin, and 10% FBS at pH 7.4, and incubated at 37°C in a humidified 5% CO 2 atmosphere.

Techniques: Inhibition, Injection, Concentration Assay, Staining

Journal: American Journal of Cancer Research

Article Title: Acid microenvironment promotes cell survival of human bone sarcoma through the activation of cIAP proteins and NF-κB pathway

doi:

Figure Lengend Snippet: Extracellular acidosis promotes cell survival in bone sarcoma via the NF-κB pathway. (A) Heat map representation of the fold increase of the expression of apoptosis and stress related-genes of osteosarcoma cells (MG63, HOS, Saos-2) after short-term acidosis. mRNA were analyzed by deep-sequencing after that cells were cultured at pH 6.5 compared to physiological medium (pH 7.4) for 24 h. Colors on the heat map indicate the log2 ratios of expression (representing normalized read counts). Red, upregulation; green, downregulation. (B) Cells were cultured under different pH (6.5 and 7.4) for 24 h as described in (A), and NF-κB mRNA expression and activation were quantified in cell lysates. (C) NF-κB1 (p50) and RelB nuclear protein concentration of osteosarcoma and Ewing sarcoma cells. Mean ± SE (n = 3 for HOS and MG63, and n = 5 for A673, *P < 0.05). (D) Signaling pathways of the tumor necrosis factor receptor (TNFR) family that mediates both survival and apoptotic pathways. The draw represents the pathways and adaptor molecules that are involved in the signaling that follows the acid stress: the NF-κB activation pathway via Akt (1), via the canonical pathway (2), via the non-canonical pathway (3), the activation of the AP1 complex transcriptional factor (4), and the TRAF-mediated death signaling with the activation of caspase cascade (5).

Article Snippet: HOS, Saos2, MG-63 (osteosarcoma), and A673 (Ewing sarcoma) human cell lines were purchased from the American Type Culture Collection, cultured in IMDM plus 20 unit/mL penicillin, 100 μg/mL streptomycin, and 10% FBS at pH 7.4, and incubated at 37°C in a humidified 5% CO 2 atmosphere.

Techniques: Expressing, Sequencing, Cell Culture, Activation Assay, Protein Concentration, Protein-Protein interactions

Journal: American Journal of Cancer Research

Article Title: Acid microenvironment promotes cell survival of human bone sarcoma through the activation of cIAP proteins and NF-κB pathway

doi:

Figure Lengend Snippet: Targeting of cIAP protein or of TRAF1 inhibit cell viability of A673 cells when cultured in acid condition. A. Inhibition of cell viability as verified by cell counting after the treatment with LCL161 (25 mM). The inhibitor was more effective at pH 6.5 than at pH 7.4. Mean ± SE (n = 3, *P < 0.05); B. Induction of apoptosis as verified by the counting of apoptotic bodies after the treatment with LCL161 (25 mM) of cells cultured at low pH (6.5). Mean ± SE (n = 3, *P < 0.05); C. mRNA analysis of TRAF1 expression by Q-RT-PCR after the treatment with specific siRNA at low pH (pH 6.5); D. Inhibition of cell viability as verified by acid phosphatase indirect assay after the treatment with specific siRNA at low pH (pH 6.5). Mean ± SE (n = 19, ****P < 0.001); E. Correlation of mRNA level between different proteins of the TRAF-cIAP destruction complex in Ewing sarcoma xenografts with acid-related markers V-ATPase V1B2 or LAMP2 (n = 6, *P < 0.05, **P < 0.01).

Article Snippet: HOS, Saos2, MG-63 (osteosarcoma), and A673 (Ewing sarcoma) human cell lines were purchased from the American Type Culture Collection, cultured in IMDM plus 20 unit/mL penicillin, 100 μg/mL streptomycin, and 10% FBS at pH 7.4, and incubated at 37°C in a humidified 5% CO 2 atmosphere.

Techniques: Cell Culture, Inhibition, Cell Counting, Expressing, Reverse Transcription Polymerase Chain Reaction