PEG-lipid-modified agonistic antibody against tumor necrosis factor receptor family elicits superior apoptosis-inducing activity against human carcinoma.

We have recently developed a novel PEG-lipid-modified antibody to enhance the induction of apoptosis by the agonistic antibody. The chemically modified TRA-8 antibody [anti-death receptor 5 (DR5) antibody] with PEG-lipid (DSPE-PEG) demonstrated significant cytotoxic activity in vitro without the need for crosslinking with a secondary antibody, which is typically required. We investigated the correlation between the PEG-lipid structure and the cytotoxic activity of the modified antibodies by varying the PEG length or lipid structure. However, when the DSPE-PEG-modified TRA-8 antibody was incubated with plasma, it lost its cytotoxic activity, likely due to degradation in the DSPE-PEG component. Nevertheless, by designing new PEG-lipids that are intended to be resistant to enzymatic degradation, we were able to prevent this degradation and restore the cytotoxic activity of the modified antibody. These findings provide valuable insights for the design of PEG-lipid-modified antibodies and suggest their potential effectiveness in enhancing cancer therapy.

Novel Immunoliposome Technology for Enhancing the Activity of the Agonistic Antibody against the Tumor Necrosis Factor Receptor Superfamily.

We have developed a technology for efficiently enhancing the anticancer apoptosis-inducing activity of agonistic antibodies against the tumor necrosis factor receptor (TNFR) superfamily by the formation of immunoliposomes. To induce apoptosis in cancer cells, agonistic antibodies to the TNFR superfamily normally need cross-linking by internal immune effector cells via the Fc region after binding to receptors on the cell membrane. To develop apoptosis-inducing antibodies that do not require the support of cross-linking by immune cells, we prepared immunoliposomes conjugated with TRA-8, an agonistic antibody against death receptor 5 (DR5), with various densities of antibody on the liposome surface, and evaluated their activities. The TRA-8 immunoliposomes exhibited apoptosis-inducing activity against various DR5-positive human carcinoma cells at a significantly lower concentration without cross-linking than that of the original TRA-8 and its natural ligand (TRAIL). The activity of the immunoliposomes was correlated with the density of antibodies on the surface. As the antibody component, not only the full-length antibody but also the Fab' fragment could be used, and the TRA-8 Fab' immunoliposomes also showed exceedingly high activity compared with the parental antibody, namely, TRA-8. Moreover, cytotoxicity of the TRA-8 full-length or Fab' immunoliposome against normal cells, such as human primary hepatocytes, was lower than that for TRAIL. Enhanced activity was also observed for immunoliposomes conjugated with other apoptosis-inducing antibodies against other receptors of the TNFR superfamily, such as death receptor 4 (DR4) and Fas. Thus, immunoliposomes are promising as a new modality that could exhibit significant activity at a low dose, for cost-effective application of an antibody fragment and with stable efficacy independent of the intratumoral environment of patients as a TNF superfamily agonistic therapy.

Fusogenic liposome can be used as an effective vaccine carrier for peptide vaccination to induce cytotoxic T lymphocyte (CTL) response.

We reported previously that fusogenic liposome (FL) introduced antigen protein encapsulated in the liposome directly into the cytoplasm of the antigen presenting cells, and that it induced immune responses. In the present study, we encapsulated TAX38-46, an HTLV-I derived protein and an antigen peptide model, into FL. The ability to induce effective cytotoxic T lymphocytes (CTL) responses in immunized mice was evaluated. Results showed FL could induce CTL response effectively and suggested that FL is a potential peptide vaccine carrier.

Augmentation of antigen-specific immune responses using DNA-fusogenic liposome vaccine.

In an attempt to enhance the immunological efficacy of genetic immunization, we investigated a new biological means for delivering antigen gene directly to the cytoplasm via membrane fusion. In this context, we investigated fusogenic liposome (FL) encapsulating DNA as a possible genetic immunization vehicle. RT-PCR analysis indicated that a FL could introduce and express encapsulating OVA gene efficiently and rapidly in vitro. Consistent with this observation, an in vitro assay showed that FL-mediated antigen-gene delivery can induce potent presentation of antigen via the MHC class I-dependent pathway. Accordingly, immunization with FL containing the OVA-gene induced potent OVA-specific Th1 and Th2 cytokine production. Additionally, OVA-specific CTL responses and antibody production were also observed in systemic compartments including the spleen, upon immunization with the OVA-gene encapsulating FL. These findings suggest that FL is an effective genetic immunization carrier system for the stimulation of antigen-specific immune responses against its encoding antigen.