cervical cancer cell line skg-iiia  (BioResource International Inc)

Journal: Molecular Therapy

Article Title: Sustainable Tumor-Suppressive Effect of iPSC-Derived Rejuvenated T Cells Targeting Cervical Cancers

doi: 10.1016/j.ymthe.2020.07.004

Figure Lengend Snippet: Generation of HPV16 E6- and E7-Specific CTLs (A) Donor characteristics and CTL epitope. (B) Flow cytometric E6 49–57 tetramer analysis of two patient donors’ lymphocytes 7 days after peptide pulse. (C) Flow cytometric E6 49–57 tetramer analysis of a healthy donor’s lymphocytes 7 days after peptide pulse (left). E6 49–57 tetramer + cells were bulk cultured (center) and single-cell cloned (right). (D) E6 49–57 tetramer + bulk-cultured CTLs were purified twice by fluorescence-activated cell sorting (FACS). (E) Flow cytometric E7 11–19 tetramer analysis of a healthy donor’s lymphocytes 7 days after peptide pulse (left). E7 11–19 tetramer + cells were bulk cultured (right). (F) E7 11-19 tetramer + bulk-cultured CTLs were purified twice by FACS. (G) Schematic illustration of establishment of T-iPSCs from HPV16 E6- and E7-CTLs.

Article Snippet: The HPV16 + cervical cancer cell lines SKG-IIIa (HLA-A24 + ), CaSki (HLA-A02 + ) (both RIKEN BioResource Research Center, Tsukuba, Ibaragi, Japan), and SiHa (HLA-A24 + ) (ATCC, Manassas, VA, USA) were cultured in RPMI 1640 medium (Sigma-Aldrich) supplemented with 10% FBS and penicillin-streptomycin-glutamine (Thermo Fisher Scientific).

Techniques: Cell Culture, Clone Assay, Purification, Fluorescence, FACS

Journal: Molecular Therapy

Article Title: Sustainable Tumor-Suppressive Effect of iPSC-Derived Rejuvenated T Cells Targeting Cervical Cancers

doi: 10.1016/j.ymthe.2020.07.004

Figure Lengend Snippet: Anti-Cervical Cancer Effect of HPV16 E6-rejTs as Examined In Vitro (A) Flow cytometric E6 49–57 tetramer analysis of Con-HPV-rejTs, 6 factors-HPV-rejTs, and bulk CTL-derived HPV-rejTs. (B) In vitro 51 Cr-release assay with HPV16 E6-rejTs (effector) and HLA-matched HPV16 + cervical cancer cell line SKG-IIIa (target). HLA-matched HPV16 − tumor cell line, control target. The effector-to-target (E:T) ratios were 10:1 and 5:1. The mean percentages of specific tumor cell lysis ± SD are shown. (C) CFSE dilution T cell proliferation assay, HPV-CTLs and HPV-rejTs. (D) Representative flow cytometric analysis of T cell subset and memory phenotype (CD45RA and CD62L population) of original HPV16 E6-CTLs and of HPV16 E6-rejTs. (E) RTCA continuous graphical output of tumor proliferation index up to 85 h for SKG-IIIa cocultured with original HPV16 E6-CTLs, 6 factors-HPV16 E6-rejTs, and control rejTs; also tumor. E:T ratios were uniformly 1:1. HLA-mismatched different epitope-rejTs, control. The data shown represent at least three independent triplicate experiments. Mean values were plotted ±SD. (F) RTCA continuous graphical output of tumor proliferation index up to 75 h for SKG-IIIa cocultured with 6 factors-HPV16 E6-rejTs, Con-HPV16 E6-rejTs, and control rejTs; also tumor only. E:T ratios were uniformly 1:1. HLA-mismatched different epitope-rejTs, control. The data shown represent at least three independent triplicate experiments. Mean values were plotted ±SD.

Article Snippet: The HPV16 + cervical cancer cell lines SKG-IIIa (HLA-A24 + ), CaSki (HLA-A02 + ) (both RIKEN BioResource Research Center, Tsukuba, Ibaragi, Japan), and SiHa (HLA-A24 + ) (ATCC, Manassas, VA, USA) were cultured in RPMI 1640 medium (Sigma-Aldrich) supplemented with 10% FBS and penicillin-streptomycin-glutamine (Thermo Fisher Scientific).

Techniques: In Vitro, Derivative Assay, Release Assay, Control, Lysis, Proliferation Assay

Journal: Molecular Therapy

Article Title: Sustainable Tumor-Suppressive Effect of iPSC-Derived Rejuvenated T Cells Targeting Cervical Cancers

doi: 10.1016/j.ymthe.2020.07.004

Figure Lengend Snippet: Anti-Cervical Cancer Effect of HPV16 E7-rejTs as Examined In Vitro and Sustainable Tumor-Suppressive Effect of HPV16 E6-rejTs In Vivo (A) Flow cytometric E7 11–19 tetramer analysis of each line of bulk CTL-derived HPV-rejTs. (B) RTCA continuous graphical output of tumor proliferation index up to 85 h for CaSki cells cocultured with HPV16 E7-rejTs, original bulk HPV16 E7-CTLs, and control CTLs; also tumor only. HLA-mismatched different epitope-CTLs, control. E:T ratios were uniformly 1:1. Data were plotted and are shown as mean ± SD. (C) Bioluminescence imaging of mice treated with original HPV16 E6-CTL clone or HPV16 E6-rejTs. GFP/FFluc -labeled cervical cancer-bearing mice were divided into three groups that received no treatment (n = 7), HPV-CTLs (n = 6), or HPV-rejTs (n = 6). Images of three representative mice from each group are shown. (D) Quantification of tumor burden on day 21 is represented. ∗p < 0.05 by one-way ANOVA. (E) Kaplan-Meier survival curves representing percentage survival of the experimental groups: no treatment, HPV-CTLs, or HPV-rejTs. ∗p < 0.05, ∗∗p < 0.01 and by log-rank testing. ns, not significant. (F) Results of comparative genomic hybridization microarray of HPV-CTLs, Con-T-iPSCs, Con-HPV-rejTs, 6 factors-T-iPSCs, and 6 factors-HPV-rejTs. No aberrations in copy numbers in all comparisons: original HPV-CTLs versus Con-T-iPSCs, Con-T-iPSCs versus Con-HPV-rejTs, original HPV-CTLs versus 6 factors-T-iPSCs, and 6 factors-T-iPSCs versus 6 factors-HPV-rejTs.

Article Snippet: The HPV16 + cervical cancer cell lines SKG-IIIa (HLA-A24 + ), CaSki (HLA-A02 + ) (both RIKEN BioResource Research Center, Tsukuba, Ibaragi, Japan), and SiHa (HLA-A24 + ) (ATCC, Manassas, VA, USA) were cultured in RPMI 1640 medium (Sigma-Aldrich) supplemented with 10% FBS and penicillin-streptomycin-glutamine (Thermo Fisher Scientific).

Techniques: In Vitro, In Vivo, Derivative Assay, Control, Imaging, Labeling, Hybridization, Microarray

Journal: International Journal of Oncology

Article Title: Eradication of cervical cancer in vivo by an AAV vector that encodes shRNA targeting human papillomavirus type 16 E6/E7

doi: 10.3892/ijo.2018.4245

Figure Lengend Snippet: Gene transfer efficiency of each AAV vector serotype in cervical cancer cells. (A) β-galactosidase expression levels at 48 h following the transduction of BOKU, SiHa and SKG-IIIa cells with the AAV vector (AAV-CMV-LacZ) types 1–9 containing the LacZ gene encoding β-galactosidase (white bars, BOKU cells; gray bars, SiHa cells; black bars, SKG-IIIa cells). The results are presented as the means ± sD. (B) gene transfer efficiency of AAV2 in cervical cancer cell lines. The GFP-positive cell rate 48 h after AAV2-shNC transduction was determined using flow cytometry.

Article Snippet: The HPV-16-positive human cervical cancer cell lines, BOKU and SKG-IIIa, and the human immortalized cell line, 293, were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan).

Techniques: Plasmid Preparation, Expressing, Transduction, Flow Cytometry

Journal: International Journal of Oncology

Article Title: Eradication of cervical cancer in vivo by an AAV vector that encodes shRNA targeting human papillomavirus type 16 E6/E7

doi: 10.3892/ijo.2018.4245

Figure Lengend Snippet: Cell growth following AAV2-shE6E7 transduction. Viable cells were measured by WST-1 assay at 72 h folllowing AAV2-shE6E7 or AAV-shNC transduction (A) BOKU cells, (B) SiHa cells, (C) SKG-IIIa cells, and (D) 293 cells; * P<0.05 as compared to the AAV-shNC transduction cells. The results are presented as the means ± SD.

Article Snippet: The HPV-16-positive human cervical cancer cell lines, BOKU and SKG-IIIa, and the human immortalized cell line, 293, were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan).

Techniques: Transduction, WST-1 Assay

Journal: International Journal of Oncology

Article Title: Eradication of cervical cancer in vivo by an AAV vector that encodes shRNA targeting human papillomavirus type 16 E6/E7

doi: 10.3892/ijo.2018.4245

Figure Lengend Snippet: In vivo tumor growth following AAV2-shE6E7 transduction. (A and B) Tumor growth curves after a single injection of AAV2-shE6E7, AAV-shNC, or saline into subcutaneous tumors with a 5-mm major axis (volume, 60 mm 3 ); (A) BOKU cells, (B) SiHa cells; * P<0.05. The results are presented as the means ± SD. (C) SiHa-derived tumors at 14 days after administration. (D and E) Tumor growth curves after a single injection of AAV2-shE6E7, AAV-shNC, or saline into subcutaneous tumors with an 8-mm major axis; (D) SiHa cells, (E) SKG-IIIa cells. (F) SiHa-derived tumors at 42 days after administration.

Article Snippet: The HPV-16-positive human cervical cancer cell lines, BOKU and SKG-IIIa, and the human immortalized cell line, 293, were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan).

Techniques: In Vivo, Transduction, Injection, Saline, Derivative Assay