Improved in vitro growth inhibitory effect of N-(phosphonacetyl)-L-aspartic acid in immunoliposomes

The use of liposome-encapsulated N-(phosphonacetyl)-L-aspartic acid (PALA) for the possible treatment of human ovarian cancer has been investigated in vitro. Protein A or tumor-specific antibodies were conjugated to liposomes via the reaction of a maleimide derivatized phospholipid (MPB-PE) with a thiol introduced into the protein by a heterobifunctional cross-linking agent, N-succimidyl 3-(2-pyridyldithio) propionate (SPDP). Antibody-conjugated PALA-containing liposomes were separated from free antibodies by ultracentrifugation in discontinuous metrizamide gradients. PALA in Protein A-conjugated liposomes was found to be over 400-fold more effective (IC50 = 0.04 mu M) than free drug (IC50 = 18 mu M) for growth inhibition of L929 cells in vitro, when the cells were pretreated with 20-40 mu g of 11-4.1 monoclonal antibody for 30 min. PALA in tumor-specific antibody-conjugated liposomes was 60-fold more effective (IC50 = 0.2 mu M) than free drug (IC50 = 12 mu M) for growth inhibition of HEY 1B human ovarian cancer cells. Anti-c-erbB2 antibody (454C11) and anti-transferrin receptor antibody (454A12) were particularly effective in this regard. For growth inhibition of SKOV-3 cells, a human ovarian cancer cell line that grows more slowly than HEY 1B, PALA in antibody-conjugated liposomes was also about 60-fold more effective (IC50 = 0.9 mu M) than free drug (IC50 = 50 mu M) Antibody against a high molecular weight glycoprotein (2G3) and anti-transferrin receptor antibody (454A12) were the most effective antibodies among those tested for their ability to inhibit growth of SKOV-3 cells. These results demonstrate that PALA is a good candidate for drug delivery to ovarian cancer cells by immunoliposomes, and that the c-erbB2 oncogene product, a high molecular weight glycoprotein, and the transferrin receptor are suitable ligands, through which to target the delivery of PALA.