Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 binds to CTLA-4 and OX40 and blocks binding to the natural ligands. ( a ) Design of ATOR-1015. The Fab domains bind to OX40. The CTLA-4 binding domains, which are fused to the light chain via a S (GGGGS) 2 linker, consists of 111 amino acids from CD86 with 5 mutations for enhanced CTLA-4 affinity. ( b ) Binding of ATOR-1015 to CTLA-4-expressing CHO cells. Cells were stained with serially diluted ATOR-1015 or IgG1 control, followed by a PE-conjugated anti-human IgG. Mean fluorescence intensity (MFI) was determined by flow cytometry ( n = 3). ( c ) ATOR-1015 completely blocks CTLA-4 from interacting with CD80 and CD86 in a competitive ELISA. Plates were coated with CD80-Fc or CD86-Fc. Serially diluted ATOR-1015 was mixed with a fixed concentration of biotinylated CTLA-4-mFc and added to the plates. Streptavidin-HRP and substrate was added, and luminescence was measured. Percent inhibition calculated based on the maximal signal in the absence of ATOR-1015 is shown ( n = 2). ( d ) Binding of ATOR-1015 to OX40 expressing CHO cells. Cells were stained as described in (B) (n = 2). ( e ) OX40 expressing CHO cells were pre-incubated with or without ATOR-1015 followed by the addition of OX40L and a fluorescently labelled detection antibody. Percent OX40L inhibition was assessed by flow cytometry (n = 3). Statistical analysis was performed using the Mann Whitney, two-tailed test (****, p < 0.0001). All data presented as mean ± SEM. n equals the number of independent experiments

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: Binding Assay, Expressing, Staining, Fluorescence, Flow Cytometry, Competitive ELISA, Concentration Assay, Inhibition, Incubation, MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 generates clustering of cells and induces T-cell activation. ( a ) ATOR-1015, the combination of monotargeting anti-CTLA-4 and anti-OX40 antibodies or IgG1 control was added to a 1:1 mix of PKH26-labelled CTLA-4-expressing HEK cells and PKH67-labelled OX40-expressing CHO cells. The formation of cell complexes was determined by flow cytometry. Data show one representative experiment out of three. ( b ) ATOR-1015 blocks CTLA-4 and promotes T-cell activation in a CTLA-4 blockade reporter assay. Jurkat T cells engineered to express CTLA-4 with an IL-2 dependent luciferase reporter were co-cultured with Raji cells expressing CD80/CD86 and an artificial T cell receptor activator. Serially diluted ATOR-1015 or IgG1 control was added. Upon addition of ATOR-1015, binding of CD80/CD86 to CTLA-4 was prevented, and co-stimulation of T cells via CD28 increased, leading to a luminescent signal. Data presented as fold induction over media control based on relative light units (RLU) (n = 2 independent experiments). ( c ) OX40-mediated T-cell activation upon ATOR-1015 crosslinking to immobilized CTLA-4. Plates were coated with CTLA-4Fc (5 μg/ml) and suboptimal levels of anti-CD3 (OKT3, 3 μg/ml). CD3 + T cells (100,000 cells/well) were added along with ATOR-1015, the combination of monotargeting antibodies and IgG1 control. After 72 h of incubation, levels of IFN-γ were measured in the supernatants by ELISA ( n = 8 donors). ( d ) OX40-mediated T-cell activation upon ATOR-1015 crosslinking to CTLA-4 on cells. HEK cells expressing CTLA-4 (30,000 cells/well) were irradiated and allowed to adhere overnight. CD4 + T cells (100,000 cells/well) were added along with anti-CD3 (UCHT1) beads, ATOR-1015, the combination of monotargeting antibodies or IgG1 control. After 72 h of incubation, levels of IL-2 were measured in the supernatants by ELISA ( n = 6 donors). ( e ) FcγRI-expressing CHO cells (100,000 cells/well) were irradiated and allowed to adhere overnight. CD4 + T cells (100,000 cells/well) were added along with anti-CD3 (OKT3) beads, ATOR-1015, the combination of monotargeting antibodies or IgG1 control. After 72 h of incubation, levels of IL-2 were measured in the supernatants by ELISA (n = 8 donors). All data presented as mean ± SEM. Statistical analysis (in C-E) was performed using the Mann Whitney, two-tailed test (**, p < 0.01; ***, p < 0.001)

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: Activation Assay, Expressing, Flow Cytometry, Reporter Assay, Luciferase, Cell Culture, Binding Assay, Incubation, Enzyme-linked Immunosorbent Assay, Irradiation, MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 induces Fcγ receptor signaling and depletion of target-expressing cells. ( a , b ) CHO cells expressing CTLA-4 and OX40 were cultured together with FcγRIIIa (F158 or V158 variant) reporter cells with an NFAT response element driving expression of a firefly luciferase. ATOR-1015, monotargeting anti-CTLA-4 and anti-OX40 antibodies alone or in combination and IgG1 control were added and Fcγ receptor activation was quantified through the luciferase produced and measured as luminescence. Data presented as fold induction based on relative light units (RLU) over media control (n = 2 independent experiments). ( c ) The expression of CTLA-4 and OX40 on the cell surface of freshly isolated and activated (stimulated for 48 h with anti-CD3/CD28 beads) Tregs determined by flow cytometry. Histogram plots from one representative donor are shown. ( d ) The expression of CTLA-4 and OX40 on the cell surface of freshly isolated and activated Tregs. Mean fluorescence intensity (MFI) was measured by flow cytometry ( n = 5 donors). ( e ) FcγRIIIa reporter cells (V158 variant) were cultured together with activated Tregs. FcγRIIIa activation was detected as described above (n = 5 donors). ( f ) Activated Tregs as target cells were cultured together with allogeneic NK cells as effector cells (effector:target cell ratio 15:1) in the absence or presence of antibodies. After 4 h, target cell lysis was measured based on lactate dehydrogenase (LDH) release ( n = 7 donors). All data presented as mean ± SEM. Statistical analysis (in D-F) was performed using the Mann Whitney, two-tailed test (**, p < 0.01; ***, p < 0.001)

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: Expressing, Cell Culture, Variant Assay, Luciferase, Activation Assay, Produced, Isolation, Flow Cytometry, Fluorescence, Lysis, MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 induces anti-tumor responses and immunological memory. Female hOX40tg mice were inoculated with MB49 cells day 0 and treated with indicated doses of ATOR-1015, monotargeting anti-CTLA-4 and anti-OX40 antibodies or vehicle day 7, 10 and 13 followed by monitoring of tumor volume and survival. ( a , b ) Tumor volume and survival of homozygous mice given vehicle or different doses of ATOR-1015 ( n = 18–19). ( c , d ) Tumor growth and survival in heterozygous mice treated with ATOR-1015, anti-CTLA-4, anti-OX40 antibodies (248 μg for bsAbs or 200 μg for mAbs) or vehicle ( n = 10). ( e ) Complete responders after treatment with ATOR-1015 as described above were re-exposed with MB49 tumor cells to demonstrate immunological memory. Naïve mice were included as tumor growth controls (n = 5). ( f ) Complete responders after treatment with ATOR-1015 as described above were re-exposed in a twin tumor model with the specific tumor (MB49) in one flank and an irrelevant tumor (PANC02) in the other to demonstrated tumor-specific memory (n = 6). All data is presented as mean ± SEM. Tumor volume was analyzed using Mann-Whitney, two-tailed test and survival using Kaplan-Meier, Log-Rank (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001). n equals the number of mice

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 localizes to the tumor and induces anti-tumor responses. ( a ) Homozygous hOX40tg mice were inoculated with MC38 cells day 0 and treated ip on days 7, 10 and 13 with antibodies (248 μg for bsAbs or 200 μg for mAbs) as indicated. Tumor volume in mice treated with ATOR-1015, surrogate anti-CTLA-4 antibodies (9D9 and 9H10) or vehicle (n = 7–10). ( b - c ) Tumor volume and survival after treatment with ATOR-1015, monotargeting anti-OX40 and anti-CTLA-4 antibodies or vehicle ( n = 25–26). ( d , e ) Mice were treated once with ATOR-1015, monotargeting anti-OX40 and anti-CTLA-4 antibodies, IgG1 control or vehicle on day 17. Twenty-four hours later, tumors and spleens were collected and the level of hIgG + cells was quantified by flow cytometry. Data show the percentage of hIgG + cells out of live CD45 + cells (n = 5–10). All data presented as mean ± SEM. Statistical differences were analyzed using Mann-Whitney, two-tailed test, except survival that was analyzed using Kaplan-Meier, Log-Rank (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001). n equals the number of mice

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: Flow Cytometry, MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 depletes Tregs and activates effector T cells in the tumor. Homozygous hOX40tg mice were inoculated with MC38 cells day 0 and treated ip with ATOR-1015, monotargeting anti-CTLA-4 and anti-OX40 antibodies (248 μg for bsAbs or 200 μg for mAbs) or vehicle on days 10, 14 and, 18. Twenty-four hours after the last injection, the tumors and spleens were harvested, and stained for Treg and effector T cell markers. ( a - b ) CD8 + T cell/Treg ratio in the tumor and spleen. ( c ) Percentage of Tregs (of CD45 + cells) in tumors. ( d ) Percentage CD8 + T cells (of CD45 + cells) in tumors. ( e - f ) Expression of CD107a and Granzyme B on CD8 + T cells in the tumors. All data presented as mean and each dot represents one animal. Statistical differences were analyzed using Mann-Whitney, two-tailed test, (*, p < 0–05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001)

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques: Injection, Staining, Expressing, MANN-WHITNEY, Two Tailed Test

Journal: Journal for Immunotherapy of Cancer

Article Title: The CTLA-4 x OX40 bispecific antibody ATOR-1015 induces anti-tumor effects through tumor-directed immune activation

doi: 10.1186/s40425-019-0570-8

Figure Lengend Snippet: ATOR-1015 improves the effect of anti-PD-1 treatment. Female heterozygous hOX40tg mice inoculated sc with MB49 tumor cells day 0 were treated ip with vehicle, 248 μg ATOR-1015, and/or 250 μg anti-PD-1 (RPM1–14) on days 7, 10 and 13. Tumor volume was measured three times per week. The number of tumor free (TF) mice in each group is indicated

Article Snippet: The ability of ATOR-1015 to block CTLA-4 and increase the co-stimulation and activation of T cells was tested in the CTLA-4 blockade reporter assay (Promega).

Techniques:

Journal: Journal of the Chinese Medical Association : JCMA

Article Title: N-acetylcysteine and atorvastatin alleviates lung injury due to ischemia-reperfusion injury in rats

doi: 10.1097/JCMA.0000000000000193

Figure Lengend Snippet: Vein gas exchange levels at the end of the experimental period. Data are presented as medians with total range. Sham (n = 6), no injury; IR (n = 6), no pretreatment; ATOR (n = 6), atorvastatin administered through gavage; NAC (n = 6), N-acetylcysteine administered intraperitoneally; and A + N (n = 6), both drugs administered. * p < 0.05 vs IR group. Vein gas pH (A); vein gas BE (B); vein gas PvCO 2 (C); vein gas HCO 3 – levels (D). A + N, atorvastatin + N-acetylcysteine; ATOR, atorvastatin; BE, base excess; HCO3-, bicarbonate; IR, ischemia-reperfusion; NAC, N-acetylcysteine; PvCO2, partial pressure of carbon dioxide; SHAM, sham treatment.

Article Snippet: The following drugs were administered as follows: NAC (Nang Kuang Pharmaceutical Co. Ltd., Tainan, Taiwan), ATOR (Pfizer Inc., New York, NY, USA), NAC + ATOR, or placebo (0.9% NaCl) for 3 days before surgery.

Techniques:

Journal: Journal of the Chinese Medical Association : JCMA

Article Title: N-acetylcysteine and atorvastatin alleviates lung injury due to ischemia-reperfusion injury in rats

doi: 10.1097/JCMA.0000000000000193

Figure Lengend Snippet: Lung injury evaluation. Representative pulmonary histological photographs of the five experimental groups. Increased leukocyte infiltration, edema, and hemorrhage were noted after IR (arrows). Pretreatment decreased the lung inflammatory infiltration. The sums of lung injury scores are presented as medians with total range. * p < 0.05 vs IR group. A + N, atorvastatin + N-acetylcysteine; ATOR, atorvastatin; IR, ischemia-reperfusion; NAC, N-acetylcysteine; SHAM, sham treatment.

Article Snippet: The following drugs were administered as follows: NAC (Nang Kuang Pharmaceutical Co. Ltd., Tainan, Taiwan), ATOR (Pfizer Inc., New York, NY, USA), NAC + ATOR, or placebo (0.9% NaCl) for 3 days before surgery.

Techniques:

Journal: Journal of the Chinese Medical Association : JCMA

Article Title: N-acetylcysteine and atorvastatin alleviates lung injury due to ischemia-reperfusion injury in rats

doi: 10.1097/JCMA.0000000000000193

Figure Lengend Snippet: Lung wet-to-dry weight ratio. Edema formation (wet/dry weight) in lung tissue increased following mesenteric ischemia-reperfusion. Data are presented as medians with total range. * p < 0.05 vs IR group. A + N, atorvastatin + N-acetylcysteine; ATOR, atorvastatin; IR, ischemia-reperfusion; NAC, N-acetylcysteine; SHAM, sham treatment.

Article Snippet: The following drugs were administered as follows: NAC (Nang Kuang Pharmaceutical Co. Ltd., Tainan, Taiwan), ATOR (Pfizer Inc., New York, NY, USA), NAC + ATOR, or placebo (0.9% NaCl) for 3 days before surgery.

Techniques:

Journal: Journal of the Chinese Medical Association : JCMA

Article Title: N-acetylcysteine and atorvastatin alleviates lung injury due to ischemia-reperfusion injury in rats

doi: 10.1097/JCMA.0000000000000193

Figure Lengend Snippet: Lung tissue cytokine levels. Lung tissue levels of TNF-α (left), IL-1β (middle), and IL-6 (right) increased following mesenteric ischemia-reperfusion. Data are presented as medians with total range. * p < 0.05 vs IR group. A + N, atorvastatin + N-acetylcysteine; ATOR, atorvastatin; IL, interleukin; IR, ischemia-reperfusion; NAC, N-acetylcysteine; SHAM, sham treatment; TNF-α, tumor necrosis factor-α.

Article Snippet: The following drugs were administered as follows: NAC (Nang Kuang Pharmaceutical Co. Ltd., Tainan, Taiwan), ATOR (Pfizer Inc., New York, NY, USA), NAC + ATOR, or placebo (0.9% NaCl) for 3 days before surgery.

Techniques:

Journal: Pharmaceuticals

Article Title: Gradient HPLC Method for Simultaneous Determination of Eight Sartan and Statin Drugs in Their Pure and Dosage Forms

doi: 10.3390/ph13020032

Figure Lengend Snippet: Application of standard addition technique for the determination of Irbesartan ® (IRB), Losazide ® (LOS), Estromap ® (ROS), Tareg ® (VAL), Ator ® (ATR), Lovastmed ® (LOV), Erastapex ® (OLM), and Alkor ® (SIM) tablets using the proposed method.

Article Snippet: , Ator ® (ATR) , , Lovastmed ® (LOV) , , Erastapex ® (OLM) , , Alkor ® (SIM) , .

Techniques: Standard Addition

Journal: Pharmaceuticals

Article Title: Gradient HPLC Method for Simultaneous Determination of Eight Sartan and Statin Drugs in Their Pure and Dosage Forms

doi: 10.3390/ph13020032

Figure Lengend Snippet: HPLC Chromatogram of authentic mixture containing Irbesartan ® (IRB), Losazide ® (LOS), Estromap ® (ROS), Tareg ® (VAL), Ator ® (ATR), Lovastmed ® (LOV), Erastapex ® (OLM), and Alkor ® (SIM) tablets dosage forms at 280 nm. Other optimum chromatographic conditions are stated in .

Article Snippet: , Ator ® (ATR) , , Lovastmed ® (LOV) , , Erastapex ® (OLM) , , Alkor ® (SIM) , .

Techniques:

Journal: Pharmaceuticals

Article Title: Gradient HPLC Method for Simultaneous Determination of Eight Sartan and Statin Drugs in Their Pure and Dosage Forms

doi: 10.3390/ph13020032

Figure Lengend Snippet: Statistical analyses of results obtained by the proposed method applied on Irbesartan ® (IRB), Losazide ® (LOS), Estromap ® (ROS), Tareg ® (VAL), Ator ® (ATR), Lovastmed ® (LOV), Erastapex ® (OLM), and Alkor ® (SIM) tablets compared with reference methods.

Article Snippet: , Ator ® (ATR) , , Lovastmed ® (LOV) , , Erastapex ® (OLM) , , Alkor ® (SIM) , .

Techniques: