Calreticulin is an effective immunologic adjuvant to tumor-associated antigens

Abstract

Abstract. As a key molecule involved in cell recognition, calreticulin (CRT) may be expressed on the surface of (pre-) apoptotic cells and provide the signal that is recognized by dendritic cells (DCs) or other antigen presenting cells (APCs), which results in phagocytosis. Within the APCs, tumor-associated antigens (TAAs) may be subsequently presented to T lymphocytes, which triggers a specific antitumor immune response. It has been hypothesized that CRT is able to act as the immunologic adjuvant and translocate itself and TAAs to the cell surface and induce a specific antitumor immune response. In the present study, CRT was demonstrated to translocate itself and mucin 1 (MUC1), a breast cancer antigen, to the surface of 4T1 cells and the MUC1-CRT-coated cells were able to induce apoptosis in a time-dependent manner. When DCs were infected with adenovirus containing MUC1-CRT, an increase in T cell proliferation and cytokine production was exhibited. These results suggest that CRT may act as an immunologic adjuvant with MUC1 and induce a strong immune response.

Silencing of suppressor of cytokine signaling 1 enhances the immunological effect of mucin 1-calreticulin-primed 4T1 cell-treated dendritic cells in breast cancer treatment

“…nos. 315-03 and 500-p45, respectively), which were used for DC cell culture as described previously ( 7 ). Anti-cluster of differentiation (CD)11c-allophycocyanin, anti-CD83-fluorescein isothiocyanate and anti-phosphorylated (p)-STAT1 antibodies were purchased from Biolegend, Inc. (San Diego, CA, USA; cat.…”

“…Bone marrow-derived dendritic cells (BMDCs) were collected and identified as previously described ( 7 ). The DC2.4 cell line was purchased from Suershengwu Co., Ltd. (Shanghai, China), the breast cancer 4T1 cell line, originating from BALB/c mice, was purchased from American Type Culture Collection (Manassas, VA, USA) and all cells were maintained in the laboratory in the China Three Gorges University.…”

“…For group A, BMDCs were incubated with 10.0 µg hpp10-DRBD and 1.0 µg SOCS1 siRNA in serum-free medium for 1 h at 37°C. For group B, pEGFP-type I transmembrane glycoprotein mucin 1 (MUC1)-CRT was constructed as previously described ( 7 ) and the primer pairs are presented in Table II . The pEGFP-MUC1-CRT plasmid (1.0 µg) was transfected into 4T1 cells to construct MUC1-CRT-primed 4T1 cells by adding 3.0 µl Lipofectamine 2000 for 4–6 h at 37°C, and the efficiency was observed using fluorescence microscopy (magnification, ×40).…”

“…BMDCs were seeded onto 6-well culture plates (1.0×10 6 /well), and splenocytes were harvested and prepared as single cell suspensions ( 7 ). Pre-treated BMDCs (as described under the ‘DC preparation’ subheading) were incubated with splenocytes at 37°C for 72 h. Following centrifugation at 800 × g for 5 min at room temperature, the supernatant was collected and culture supernatants (100 µl/well) were used to analyze production of the cytokines IFN-γ and TNF-α by ELISA, according to the manufacturer's protocol.…”

“…In cancer immunotherapy, it is essential to sufficiently expose tumor-associated antigens (TAAs) to DCs in order to stimulate DC maturation and facilitate the subsequent activation of antigen-specific immune responses. Our previous data revealed that calreticulin (CRT) is a key molecule involved in cell recognition and that it may act as an ‘immunological adjuvant’ by translocating type I transmembrane glycoprotein mucin 1 (MUC1), a breast cancer antigen, to the surface of 4T1 cells ( 7 ). The MUC1-CRT-primed 4T1 cells were recognized by DCs, leading to a specific antitumor immune response.…”