Healthy B cells become susceptible to APECs following the provision of exogenous MMP2 protease. MMP2-cleavable cetuximab-APECs are killed by peptide-specific CMV-CTLs. An MMP2-cleavable cetuximab APEC was used to label the breast cancer cell line MDA-MB-231. After co-incubation of labeled tumor cells with CMV-CTLs, tumor cell lysis occurred as T cells recognized the CMV epitope being presented at the cell surface by tumor cells. NIHMS1612332-supplement-Suppl_Video_5_Breast_cancer_cells_labeled_with_an_MMP2-cleavable_cetuximab-APECs_are_killed_by_peptide-specific_CMV-CTLs__An_MMP2-cleavable_cetuximab_APEC_was_used_to_label_the_breast_cancer_cell_line_MDA-MB-231__After.mp4 (30M) GUID:?94BA240C-0BCD-490B-9471-83AA1E6050F9 Suppl.Video 3:Cetuximab-labeled breast cancer cells are not recognized by CMV-CTLs. Breast cancer cells (MDA-MB-231) were labeled with the anti-EGF receptor antibody cetuximab and incubated with CMV-CTLs. There was no recognition of tumor cells by CMV-CTLs: Suppl.Video 3:Cetuximab-labeled breast cancer cells are not recognized by CMV-CTLs. Breast cancer cells (MDA-MB-231) were labeled with the anti-EGF receptor antibody cetuximab and incubated with CMV-CTLs. There was no recognition of tumor cells by CMV-CTLs. NIHMS1612332-supplement-Suppl_Video_3_Cetuximab-labeled_breast_cancer_cells_are_not_recognized_by_CMV-CTLs__Breast_cancer_cells__MDA-MB-231__were_labeled_with_the_anti-EGF_receptor_antibody_cetuximab_and_incubated_with_CMV-CTLs__There_was_no_.mp4 (32M) GUID:?6CA1AC99-C335-4252-865D-6FE806C1281C Suppl.Video 1Untreated breast cancer cells do not instigate CMV-CTL cytotoxicity. Real-time imaging of MDA-MB-231 cell line containing a red nuclear dye to determine cytotoxicity by NLV-specific CMV-CTLs over 40?h. Untreated MDA-MB-231 cells were not targeted by CMV-CTLs during co-culture: Suppl.Video 1Untreated breast cancer cells do not instigate CMV-CTL cytotoxicity. Real-time imaging of MDA-MB-231 cell line containing a red nuclear dye to determine cytotoxicity by NLV-specific Chlorzoxazone CMV-CTLs over 40 h. Untreated MDA-MB-231 cells were not targeted by CMV-CTLs during co-culture. NIHMS1612332-supplement-Suppl_Video_1Untreated_breast_cancer_cells_do_not_instigate_CMV-CTL_cytotoxicity__Real-time_imaging_of_MDA-MB-231_cell_line_containing_a_red_nuclear_dye_to_determine_cytotoxicity_by_NLV-specific_CMV-CTLs_over_40_h__Unt.mp4 (33M) GUID:?3CD09EBE-D1DF-4E96-ABE7-BF439049FE52 Suppl.Video 2:Breast cancer cells exogenously labeled with CMV peptide are killed by peptide-specific CMV-CTL. Breast cancer cells (MDA-MB-231) were exogenously labeled with CMV peptide (NLVPMVATV) and co-incubated with peptide-specific CMV-CTLs. During co-culture, CMV-CTLs recognized the peptide displayed by the tumor cells and efficiently killed the breast cancer cells: Suppl.Video 2:Breast cancer cells exogenously labeled with CMV peptide are killed by peptide-specific CMV-CTL. Breast cancer cells (MDA-MB-231) were exogenously labeled Chlorzoxazone with CMV peptide (NLVPMVATV) and co-incubated with peptide-specific CMV-CTLs. During co-culture, CMV-CTLs recognized the peptide displayed by the tumor cells and efficiently killed the breast cancer cells. NIHMS1612332-supplement-Suppl_Video_2_Breast_cancer_cells_exogenously_labeled_with_CMV_peptide_are_killed_by_peptide-specific_CMV-CTL__Breast_cancer_cells__MDA-MB-231__were_exogenously_labeled_with_CMV_peptide__NLVPMVATV__and_co-incubated_wit.mp4 (29M) GUID:?AD108569-9ACB-4C78-AF27-0322AB8D0708 Suppl.Video 4:Negative control cetuximab-APEC-labeled breast cancer cells are not recognized by CMV-CTLs. A negative control cetuximab APEC, designed to prevent release of the CMV epitope, was used to label the breast cancer cell line MDA-MB-231. After co-incubation of labeled tumor cells with CMV-CTLs, tumor cells were not targeted for killing by T cells: Suppl.Video 4:Negative control cetuximab-APEC-labeled breast cancer cells are not recognized by CMV-CTLs. A negative control cetuximab APEC, designed to prevent release of the CMV epitope, was used to label the breast cancer cell line MDA-MB-231. After co-incubation of labeled tumor cells with CMV-CTLs, tumor cells were not FANCG targeted for killing by T cells. NIHMS1612332-supplement-Suppl_Video_4_Negative_control_cetuximab-APEC-labeled_breast_cancer_cells_are_not_recognized_by_CMV-CTLs__A_negative_control_cetuximab_APEC__designed_to_prevent_release_of_the_CMV_epitope__was_used_to_label_the_breast_.mp4 (30M) GUID:?F86EF398-32BA-4B56-9044-96CC2A053300 Suppl.Tables 1-4. NIHMS1612332-supplement-Suppl_Tables_1-4.pdf (1.7M) GUID:?803CF476-5B66-4941-9E64-5DB060F17B1E Data Availability StatementData presented in this study are available in the article, Supplementary Information or from the corresponding author on reasonable request. Abstract Several cancer immunotherapy approaches, such as immune checkpoint blockade and adoptive T-cell therapy, boost T-cell activity against the tumor, but these strategies are not effective in the absence of T cells specific for displayed tumor antigens. Here we outline Chlorzoxazone an immunotherapy in which endogenous T cells specific for a noncancer antigen are retargeted to attack tumors. The approach relies on the use of antibody- peptide epitope conjugates (APECs) to deliver suitable antigens to the tumor surface for presention by HLA-I. To Chlorzoxazone retarget cytomegalovirus (CMV)-specific CD8+ T cells against tumors, we used APECs containing CMV-derived epitopes conjugated to tumor-targeting antibodies via metalloprotease-sensitive linkers. These APECs redirect pre-existing CMV immunity against tumor cells in vitro and in mouse cancer models. In vitro, APECs activated specifically CMV-reactive effector T cells whereas a bispecific T-cell engager activated both.