Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: Tumor-associated antigen expression (normalized MFI, mean fluorescence intensity) in PDAC cell lines

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: Expressing, Fluorescence

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: Structure and in vitro activity of PDAC-directed T cell-engaging bispecific antibodies ( A ) IgG-[L]-scFv structure of BsAbs. CH, constant heavy chain; CL, constant light chain; scFv, single chain variable fragment; VH, variable heavy chain; VL, variabl light chain. ( B ) Flow cytometric analysis displaying fluorescent intensities of IgG-[L]-scFv BsAbs bound to human PDAC cells. IgG-[L]-scFv BsAbs evaluated were specific for human EGFR, HER2, MSLN, c-MET, and B7-H3. ( C ) Antibody-dependent T cell-mediated cytotoxicity (ADTC) as a function of increasing doses of BsAbs against PDAC cell lines. Ratio of effector T cells to target PDAC cells (E: T ratio) was set to 10:1

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: In Vitro, Activity Assay

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: In vitro sensitivities (EC50, pM) to target antigen-specific bispecific antibodies in PDAC cell lines

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: In Vitro

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: EGFR and HER2 T-BsAb heterodimerization and in vitro kinetics ( A ) Schematic of BsAb heterodimerization by controlled Fab Arm Exchange. Parental BsAbs bore 2 Fabs specific to EGFR or HER2 and 2 scFvs specific for CD3 (‘2 + 2’). Reduction of inter-H-chain disulfide bonds and subsequent heterodimerization (driven by F405L and K409R amino acid substitutions) yielded EGFRxHER2 BsAbs bearing 1 EGFR Fab, 1 HER2 Fab, and 2 CD3 scFv (‘1 + 1 + 2’). ( B ) SPR analysis of T-BsAbs binding to EGFR (left panel), HER2 (middle panel), and EGFR x HER2 (right panel) proteins. Representative normalized sensorgrams at 20nM were shown. ( C ) IHC staining of two cell-derived PDAC xenograft tumors SW1990 (top) and BxPC-3 (bottom). OCT-embedded tumor sections were stained with EGFRxEGFR, HER2xHER2, or EGFRxHER2 BsAbs. Control slides were either unstained (SW1990) or incubated with a control antibody (BxPC-3). ( D ) Antibody-dependent T cell-mediated cytotoxicity assays (ADTC) against SW1990 (left) and BxPC-3 (right). Cytotoxicity measured by Chromium-51 release. Ratio of effector T cells to target PDAC cells (E: T ratio) was set to 10:1. EC50s (SW1990; BxPC-3): EGFRxEGFR: 165.8fM; 31.5fM, HER2xHER2: 18.5pM; 7.14pM, EGFRxHER2: 699.6fM; 128.8fM. CD33xCD33 BsAb was included as a negative control

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: In Vitro, Binding Assay, Immunohistochemistry, Derivative Assay, Staining, Control, Incubation, Negative Control

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: Avidities of EGFR and HER2 T-BsAbs

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: Protein Binding

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: Heterodimeric EGFR and HER2 T-BsAbs impede PDAC growth ( A ) In vivo antitumor effect of BsAbs in the presence of human T cells against SW1990 cell line xenografts; 3 × 10 6 cells of SW1990 were subcutaneously implanted into mice. T-BsAbs were administered twice per week and 2 × 10 7 of luciferase-transduced T cells (Luc(+) T cells) were administered once per week for 2 weeks to treat the tumors. ( B ) In vivo anti-tumor effects of 10 µg EGFR and HER2 T-BsAbs. ( C ) Relative body weight of mice during treatment and ( D ) overall survival were plotted. ( E ) Bioluminescence imaging (BLI) of Luc(+) T cells. Quantitation of bioluminescence intensity in the lesions of tumors. Representative images (right) were taken on day 7

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: In Vivo, Luciferase, Imaging, Quantitation Assay

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: T-BsAbs drive T cell infiltration, activation, and function in PDAC ( A ) Treatment of SW1990 tumor-bearing mice with EGFR and HER2 T-BsAbs; 3 × 10 6 cells of SW1990 were subcutaneously implanted into mice. Once tumor reached ∼500mm 3 , mice received a single infusion of 2 × 10 7 T cells and were treated with 10µg T-BsAb, administered twice per week. Tumors were harvested for analysis on day 10 after initiation of treatment. Mice that exhibited 50% or more reduction in tumor volume by day 10 were excluded from this and subsequent analyses. ( B ) Flow cytometric analysis of the in vivo effect of T-BsAb treatments on T cell infiltration into harvested SW1990 PDAC tumors. Infiltrating T cells were identified by detection of human CD45 + and human CD4 + or CD8 + . ( C ) Frequency of T cell activation indicated by detection of IFN-γ + TNF-α + T cells and expression of CD69 among CD4 + or CD8 + T cells. ( D ) Frequency of CD11b + intratumoral myeloid cells and prevalence of PDL1 expression

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: Activation Assay, In Vivo, Expressing

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: ADTC sensitivities (EC50, pM) of SW1990 lines to EGFR and HER2 T-BsAbs

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques:

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: FACS binding (EC50, pM) of EGFR and HER2 T-BsAbs to SW1990 lines

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: Binding Assay

Journal: Journal of Hematology & Oncology

Article Title: Heterodimerization of T cell engaging bispecific antibodies to enhance specificity against pancreatic ductal adenocarcinoma

doi: 10.1186/s13045-024-01538-5

Figure Lengend Snippet: EGFRxHER2 T-BsAbs efficacy is lost in EGFR and HER2-single-positive PDAC ( A ) Mean fluorescent intensities determined by flow cytometry and ( B ) antibody-dependent T cell-mediated cytotoxicity (ADTC) against SW1990-EGFR-KO (left) and SW1990-HER2-KO (right) cell lines. Ratio of effector T cells to target PDAC cells (E: T ratio) was set to 10:1. ( C ) Tumor growth of subcutaneous SW1990 EGFR-KO (left) and HER2-KO (right) tumors. One infusion of 2 × 10 7 T cells was administered. Two doses of T-BsAbs (10 µg each) were given by i.v. injection

Article Snippet: Either recombinant human EGFR (R&D system: Cat# 9565-ER), HER2 (Sino Biological: Cat# 10,004-H08H), or both were immobilized on CM5 chips.

Techniques: Flow Cytometry, Injection

Journal: Cancers

Article Title: Drug-Induced Resistance and Phenotypic Switch in Triple-Negative Breast Cancer Can Be Controlled via Resolution and Targeting of Individualized Signaling Signatures

doi: 10.3390/cancers13195009

Figure Lengend Snippet: TNBC tissues are represented by different patient-specific signaling signatures, majority of which do not include EGFR. ( A ) Fold changes in expression levels of EGFR and pEGFR in TNBC and non-TNBC tumors are shown. Y1068 and Y1173 are EGFR phosphorylation sites; ( B ) Examples for patient-specific sets of active unbalanced processes are shown. Each sample harbors a set of 1–3 active unbalanced processes (PaSSS), represented schematically by a barcode. In each barcode active unbalanced processes are represented by black or gray squares, inactive white. Negative/positive amplitude denotes how the patients are correlated with respect to a particular process. Suggested PaSSS-based therapies appear below each barcode; ( C ) Heterogeneity index of 3 subgroups, represented by a ratio between the number of distinct PaSSSs and the number of samples in each subset, is shown for the TNBC subset of tissues, the entire set (3467 samples from 11 cancer types) and the subset of non-TNBC samples. (Abbreviations: TNBC—Triple Negative Breast Cancer, PaSSS—Patient-specific signaling signature, EGFR—Epidermal Growth Factor Receptor, VEGFR2—Vascular Endothelial Growth Factor Receptor 2, Her2—Human Epidermal growth factor Receptor 2, Src—Proto-oncogene tyrosine-protein kinase Src).

Article Snippet: The following conjugated antibodies were used: anti-p-EGFR (Y1068) (R&D Systems, Minneapolis, MN, USA, cat. no. IC3570G), anti-p-ERK2 (Thr202/Tyr204) (BioLegend, San Diego, CA, USA, cat. No. 675503), anti-p-S6 (Ser235/236) (BioLegend, cat. no. 608605), and anti-GAPDH (Santa Cruz Biotechnology, Dallas, Texas, USA, cat. no. sc-47724AF594).

Techniques: Expressing, Phospho-proteomics