A new purification method for enhancing the immunogenicity of heat shock protein 70-peptide complexes

Abstract

When purified from a tumor, certain heat shock protein 70 (HSP70)-peptide complexes (PCs) can function as effective vaccines against the tumor from which the complexes were isolated. The immunogenic mechanisms of HSP70 preparations imply that tumor-derived HSP70-PCs exhibit antigens associated with antigen-presenting cells such as dendritic cells (DCs), inducing antigen-specific cytotoxic CD8+ T cells. However, some important membrane-resident tumor-associated peptides, such as the HER-2/neu (c-erbB2) oncogenic protein, cannot be purified from HSP70 by traditional methods. In the present study, a new approach for the purification of HSP70-PCs from HER-2-overexpressing breast cancer cells was established. The detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) was used to obtain more effectual tumor peptides. The new purified product was named HSP70-HER-2-PC, and its immunological activities were determined. Traditionally purified HSP70-PCs (without CHAPS) and recombinant human HSP70-HER-2 protein complexes (recombined in vitro) were used as controls. These three HSP70-associated tumor antigenic complex pulsed dendritic cells (DCs) were used to stimulate an antitumor response. The mature DCs pulsed with HSP70-HER-2-PCs stimulated autologous T cells to secrete higher levels of type I cytokine compared to the two control groups. Moreover, DCs pulsed with HSP70-HER-2-PCs induced the most specific CD8+ T cells that specifically killed the same tumor cells. These findings provide a basis for new approaches in enhancing HSP70-based immunotherapy for HER-2-associated or other membrane antigenic peptide-related cancers.

Enhancing the treatment effect on melanoma by heat shock protein 70-peptide complexes purified from human melanoma cell lines

“…Early studies revealed many important antigens in human tumor cell lines. They used lysates of human tumor cell lines to pulse DCs for treatment of the same kind of tumor and achieved a certain therapeutic effect ( 4 , 12 , 13 , 24 , 25 ).…”

“…Our previous study revealed that HSP70-PCs purified from tumor cells have better immunocompetence than lysates of the same cells. Moreover, we established a new method using the detergent CHAPS to purify HSP70-PCs containing more efficient tumor peptides from human cancer cells ( 12 ).…”

“…The methods used to purify HSP70-PCs have been described previously ( 12 ). Human melanoma cell lines were heated in a water bath at 42°C for 12 h, followed by recovery for 2 h at 37°C in an atmosphere containing 5% CO 2 .…”

“…Heat shock protein 70 (HSP70) has an important role as a molecular chaperone that binds to tumor antigen peptides in tumor cells ( 9 – 11 ). In our previous study, we purified HSP70-HER-2-peptide complexes (PCs) from the human breast cancer cell line SKBR-3, and showed that HSP70-HER-2-PCs contain more comprehensive tumor antigen peptides and induce stronger immune activity ( 12 ). Numerous studies show that human melanoma cell lines also contain a number of effective tumor antigen peptides ( 13 , 14 ).…”

Dendritic-Tumor Fusion Cells Derived Heat Shock Protein70-Peptide Complex Has Enhanced Immunogenicity

“…Some alternatives have been tested, including HSPs-pulsed dendritic cells[ 15 ], chaperone-rich cell lysates derived from tumors(CRCL) [ 23 ], combination with GM-CSF [ 18 , 24 ], recombinant HSPs with tumor-associated peptides[ 25 ] or encapsulated in nanoemulsions [ 26 ] and have shown improved immunogenicity. Furthermore, improved antitumor immune responses have been observed following the development of specific new methods of vaccine preparation for enhanced immunogenicity[ 27 , 28 ].…”