A Crucial Cytotoxic Role of Anti-Idiotypic Antibody in Immunotherapy for B-Cell Neoplasms with Tumor Cell-Derived Heat Shock Protein 70.

Tumor-derived heat shock proteins (HSPs), which bind to tumor-specific antigenic peptides, can be used for cancer immunotherapy. In this meeting, we previously reported that vaccination with mouse B-cell leukemia/lymphoma cell line, A20, -derived HSP70 (A20-HSP) against A20 cells in mice induces tumor-specific cellular immune responses (Sato et al. Blood 2001;98:1852-1857; Iuchi et al. Int J Hematol 2006;84:449-458) as well as A20-specific humoral immunity through complement dependent cell-cytotoxicities (CDCs) (Sato K et al., 2007, Blood 110:682a, Abstruct#2302). In the B-cell neoplasms, idiotype is known as one of the important tumor specific antigens which induce anti-idiotype cellular and humoral immune responses. A20-secreted monoclonal IgG (A20-Ig) and A20-idiotypic epitope peptide (A20-IP: DYWGQGTEL), which is derived from variable region of the heavy chain of the A20-Ig, are known as the A20-specific antigens (J Immunol. 2002; 168: 3983-91). Further analysis using A20-Ig and A20-IP enables to establish novel HSP70-based therapeutic strategies for B-cell neoplasms. The present study investigated whether immunization with A20-HSP induces anti-idiotypic antibodies, and also evaluated whether the antibodies showed CDCs.

A20 and syngeneic Balb/c mice (H-2K^d) were used in this study. HSP70 was purified from either A20 cells or healthy mouse liver tissue. A20-Ig was purified from A20 culture supernatants. After the subcutaneous immunization with A20-HSP, liver-derived HSP70 (control) to the healthy mice, the sera were harvested for the following experiments. To detect anti-A20-IP antibodies, the sera were assayed by ELISA to detect the specific IgG against A20-HSP, or A20-Ig. To confirm that immunization with A20-HSP induces anti-idiotypic antibodies, we analyzed the inhibitory effect (% inhibition) of A20-IP on the A20-HSP-mice sera reactivity against A20-HSP by preincubation of the sera with A20-IP or H-2K^d biding control peptide (mouse influenza hemagglutinin peptide: IYSTVASSL) by ELISA. To confirm that A20-HSP mice sera contains CDC type antibody directed to A20-IP, the CDC activity was determined by the trypan blue uptake of A20 cells after incubation with the complement and A20-HSP mice sera preincubated without or with A20-IP or control peptide.

The IgG level in the A20-HSP mice against either A20-HSP or A20-Ig was significantly increased in comparison to that in control mice. % inhibition of A20-IP (45.7%) was significantly higher than that of control peptide (1.0%), indicating that almost half of IgG in the A20-HSP mice sera which reacts to A20-HSP recognizes A20-IP. The CDC activity of A20-HSP mice sera against A20 was markedly inhibited by preincubation of the sera with A20-IP but with control peptide. Conclusions: Immunization with A20-HSP70 induces anti-idiotype antibody and the antibody contributes a crucial role in eradication of A20 by CDC activity in mice. These findings enable to establish a novel and advantageous therapeutic strategy against B-cell neoplasms utilizing the anti-idiotypic antibody in HSP-based autologous tumor immunotherapy.

No relevant conflicts of interest to declare.