Journal: Molecular Therapy Oncology

Article Title: KLS-3021: Innovative oncolytic virotherapy for the treatment of advanced primary and metastatic cutaneous squamous cell carcinoma

doi: 10.1016/j.omton.2026.201182

Figure Lengend Snippet: In vitro cytotoxicity and tumor-restricted replication analysis of KLS-3021 in cSCC cells (A) Schematic representation of KLS-3021 showing deletions of C11R , K3L , and J2R loci and insertions of therapeutic genes encoding PH-20, IL-12, and sPD1-Fc. (B) Cytotoxicity of KLS-3021 in human cSCC cell lines (A-431 and HSC-1) and normal human epidermal keratinocytes (NHEKs) was measured via CCK-8 assay at 72 h post-infection. (C) Viral replication in cSCC cell lines and NHEKs was determined via TCID 50 assay at 72 h post-infection. Data was represented as mean ± SEM of three independent experiments.

Article Snippet: Luciferase-labeled A-431 cells were cultured in DMEM (ATCC) supplemented with 10% FBS (Gibco), 1% penicillin-streptomycin (Sigma), and 8 μg/mL blasticidin (InvivoGen).

Techniques: In Vitro, CCK-8 Assay, Infection

Journal: Molecular Therapy Oncology

Article Title: KLS-3021: Innovative oncolytic virotherapy for the treatment of advanced primary and metastatic cutaneous squamous cell carcinoma

doi: 10.1016/j.omton.2026.201182

Figure Lengend Snippet: Tumor growth analysis in orthotopic primary cSCC model following KLS-3021 treatment (A) Schematic of the experimental design for the orthotopic cSCC model established by intradermal implantation of A-431 cells. (B) Tumor growth curves of vehicle-treated and KLS-3021-treated mice (mean ± SEM, n = 6 per group; ∗∗∗ p < 0.001 by two-tailed t test). (C) Individual tumor growth curves for each mouse. (D) Body-weight changes during the observation period were represented (mean ± SEM). (E and F) Representative hematoxylin and eosin (H&E)-stained sections of tumors collected on days 3 (E) and 7 (F) post-treatment. Scale bars, 300 μm; enlarged views, 100 μm.

Article Snippet: Luciferase-labeled A-431 cells were cultured in DMEM (ATCC) supplemented with 10% FBS (Gibco), 1% penicillin-streptomycin (Sigma), and 8 μg/mL blasticidin (InvivoGen).

Techniques: Two Tailed Test, Staining

Journal: Molecular Therapy Oncology

Article Title: KLS-3021: Innovative oncolytic virotherapy for the treatment of advanced primary and metastatic cutaneous squamous cell carcinoma

doi: 10.1016/j.omton.2026.201182

Figure Lengend Snippet: Bioluminescence imaging and tissue analysis in metastatic cSCC model following KLS-3021 treatment (A) Schematic of the experimental design for the orthotopic metastatic cSCC model established by intradermal implantation of luciferase-labeled A-431 cells into the footpad. (B) Longitudinal quantification of total photon flux from the primary tumor and popliteal lymph node (PLN) at each time point post-treatment ( n = 5 per group; ∗∗∗ p < 0.001 by two-tailed t test). (C) Representative IVIS bioluminescence images showing signals from primary and nodal tumors in vehicle- and KLS-3021-treated mice at indicated time points. (D and E) Primary tumor weights (D) and PLN weights (E) measured on day 3 ( n = 5 per group; ∗ p < 0.05 by two-tailed t test) and day 28 ( n = 10 per group; ∗ p < 0.05, ∗∗∗ p < 0.001 by two-tailed t test) post-treatment. All data were represented as mean ± SEM.

Article Snippet: Luciferase-labeled A-431 cells were cultured in DMEM (ATCC) supplemented with 10% FBS (Gibco), 1% penicillin-streptomycin (Sigma), and 8 μg/mL blasticidin (InvivoGen).

Techniques: Imaging, Luciferase, Labeling, Two Tailed Test

Journal: Science Advances

Article Title: Intracellular enzymatic reducing systems control receptor tyrosine kinase signaling via PTP1B

doi: 10.1126/sciadv.adv5362

Figure Lengend Snippet: ( A ) A431 cells overexpressing GLRX1 or GLRX2, indicated with upward arrows, were serum starved before stimulation with EGF (100 ng/ml) for 0-2-4-6 min. Lysates were analyzed with immunoblots (IB) for total EGFR, for total phosphotyrosine (total pTyr), for the preferred site for PTP1B-mediated dephosphorylation (pY992), and for GLRX1 or GLRX2 expression levels, as indicated. ( B and C ) Quantifications using densitometry of total pTyr and pTyr992, respectively. The signals were normalized for the intensity of the total expression of EGFR and compared with signals in controls. ( D ) GSH levels were determined in A431 cells with or without overnight treatment with 125 μM buthionine sulfoximine (BSO), as indicated. EGF stimulation and GLRX1 or GLRX2 overexpression as in (A). ( E to G ) Immunoblots and quantification of total pTyr and pTyr992 in the cells analyzed in (D). The signals were normalized for the intensity of the total expression EGFR. EV, cells transfected with control vector; GLRX1↑ and GLRX2↑, cells transfected for overexpression of GLRX1 or GLRX2, respectively. Bar graphs are determinations from immunoblots of three separate cell experiments [ n = 3; means ± SE (error bars); * P < 0.05]. Data were analyzed using one-way analysis of variance followed by Bonferroni post hoc tests for multiple comparisons with GraphPad Prism.

Article Snippet: The epidermoid carcinoma cell line A431 (American Type Culture Collection) or MEFs were cultured in DMEM (5% CO 2 ) + 10% (v/v) fetal bovine serum (FBS), 2 mM l -glutamine, penicillin, and streptomycin.

Techniques: Western Blot, De-Phosphorylation Assay, Expressing, Over Expression, Transfection, Control, Plasmid Preparation

Journal: Science Advances

Article Title: Intracellular enzymatic reducing systems control receptor tyrosine kinase signaling via PTP1B

doi: 10.1126/sciadv.adv5362

Figure Lengend Snippet: A431 cells with knockdown of ( A ) GLRX2 or ( F ) TXN1, indicated with downward arrows, were serum starved before stimulation with EGF (100 ng/ml) for 4 min, after which EGFR receptor inhibitor (10 μM) was added. Cells were harvested at indicated time points, and receptor phosphorylation (total pTyr and pY992) was analyzed by immunoblotting. ( B , C , D , and E ) The densitometric signals were normalized for the intensity of the total expression of the EGFR. The same samples were used to confirm knockdown of TXN1 and GLRX2. Quantifications and densitometry of total pTyr and pTyr992 in respective experiments as shown in (A) and (D). The signals were normalized for the intensity of the total expression of the EGFR at 4 min. ( G and H ) Analyses of PRDX1, PRDX2 or PRDX3 dimer form versus total ratio for each PRDX isoform, in arbitrary units (Dimer/tot AU) after GLRX1, GLRX2, or TXN1 knockdown, as indicated. Bar graphs are determinations from immunoblots of three separate cell experiments [ n = 3; means ± SE (error bars); * P < 0.05]. ( I , J , and K ) A431 cells with or without knockdown of GLRX2 or overexpression of GLRX2 were serum starved before stimulation with EGF (100 ng/ml). At the indicated times after EGF stimulation, cells were subjected to the modified cysteinyl-labeling assay using biotinylated iodoacetyl-PEG2-biotin for analysis of reversible PTP1B oxidation. Biotinylated proteins were purified using streptavidin-Sepharose beads and resolved by SDS-PAGE as illustrated in the scheme (K). Visualization was performed using antibodies against PTP1B, and control levels were determined from total cell lysate by SDS-PAGE and blotting against PTP1B. The densitometric signals were normalized for the intensity of PTP1B streptavidin-biotin signals over total amount of PTP1B, thus indicating the ratio of oxidized PTPB1B over total PTP1B (oxPTP1B/Total PTP1B), in arbitrary units (a.u.). Graphs are determinations from immunoblots of three (GLRX2) separate experiments [ n = 4; means ± SE (error bars); * P < 0.05]. Data were analyzed using one-way analysis of variance followed by Bonferroni post hoc tests for multiple comparisons with GraphPad Prism.

Article Snippet: The epidermoid carcinoma cell line A431 (American Type Culture Collection) or MEFs were cultured in DMEM (5% CO 2 ) + 10% (v/v) fetal bovine serum (FBS), 2 mM l -glutamine, penicillin, and streptomycin.

Techniques: Knockdown, Phospho-proteomics, Western Blot, Expressing, Over Expression, Modification, Labeling, Purification, SDS Page, Control

Journal: Science Advances

Article Title: Intracellular enzymatic reducing systems control receptor tyrosine kinase signaling via PTP1B

doi: 10.1126/sciadv.adv5362

Figure Lengend Snippet: A431 cells were starved for overnight, treated with EGF (100 ng/ml) for 2 and 4 min and subsequently with 10 μM EGFR inhibitor for 30 s, 2 min, and 6 min, as indicated. The cells were fixed and processed for PLA to determine protein interactions. Cells were stained with primary antibodies from different species against PTP1B (mouse) together with antibodies against ( A ) GLRX1 (goat), ( B ) GLRX2 (rabbit), ( C ) TXN1 (rabbit), or ( D ) EGFR (goat) to reveal protein-protein interaction in situ using the technique of PLA. The cells were fixed and processed for PLA to determine the direct interactions. Red spots indicate protein interactions with nuclei counterstained with DAPI (blue). ( E ) Bar graphs with quantifications and statistics of PLA signals (** P < 0.01; **** P < 0.0005); n ≥ 3 (more than 500 cells per experiment). Error bars denote SEM. Scale bar, 30 μm. Data were analyzed using one-way analysis of variance followed by Bonferroni post hoc tests for multiple comparisons with GraphPad Prism.

Article Snippet: The epidermoid carcinoma cell line A431 (American Type Culture Collection) or MEFs were cultured in DMEM (5% CO 2 ) + 10% (v/v) fetal bovine serum (FBS), 2 mM l -glutamine, penicillin, and streptomycin.

Techniques: Staining, In Situ