ABSTRACT
OBJECTIVES: The plasminogen activator inhibitor type 2 (PAI2) when labelled with (213)Bi forms the (213)Bi-PAI2 alpha conjugate (AC). This AC has been shown to be efficacious in preclinical studies with breast, ovarian, prostate and pancreatic cancers. The objectives of this study were to investigate the pharmacokinetics and in vivo stability of (213)Bi-PAI2 in mice, its toxicity in mice and rabbits; and to determine whether a prior injection of a metal chelator (Ca-DTPA) or lysine can reduce toxicity by decreasing renal uptake. METHODS: Two chelators (CHX-A"-DTPA and cDTPA) were used for preparation of the (213)Bi-PAI2 conjugate, for intraperitoneal administration in mice and ear vein injection in rabbits. The mice were sacrificed at different time points for pharmacokinetic studies. Blood and organs were collected for toxicity studies for all groups. RESULTS: Both chelators were found to have similar %ID/g in the kidneys over four hours. Mice and rabbits did not show any short term toxicity over 13 weeks at 1420 MBq/kg and 120 MBq/kg (213)Bi-PAI2 respectively. Kidney uptake was decreased three fold by lysine. Radiation nephropathy was observed at 20-30 weeks in mice, leading to severe weight loss, whereas severe and widespread renal tubular necrosis was observed at 13 weeks in rabbits. CONCLUSIONS: Radiation nephropathy is the dose limiting toxicity observed in mice and rabbits. Lysine can reduce kidney uptake by three fold. Based on long-term monitoring, the maximum tolerance doses (MTD) are 350 and 120 MBq/kg for mice and rabbits respectively.
Subject(s)
Bismuth/pharmacokinetics , Bismuth/toxicity , Plasminogen Activator Inhibitor 2/pharmacokinetics , Plasminogen Activator Inhibitor 2/toxicity , Radioimmunotherapy/methods , Radioisotopes/toxicity , Alpha Particles , Animals , Chelating Agents/pharmacology , Female , Kidney/pathology , Medical Oncology/methods , Mice , Mice, Inbred BALB C , Neoplasm Metastasis , Rabbits , Radioisotopes/chemistry , Time FactorsABSTRACT
Metastasis is the principal cause of death in breast cancer patients. New and improved treatments for eradicating micrometastases are needed. To this end, a novel alpha-emitting protein construct, 213Bi-labelled plasminogen activator inhibitor type-2 (PAI-2) (alpha-PAI-2), was evaluated in vitro. This construct exploits: (a) the overexpression of the cell-surface receptor bound urokinase plasminogen activator (uPA) in the metastatic spread of breast cancer cells; (b) the binding and inhibition of receptor-bound uPA by PAI-2; and (c) the high cytotoxicity of alpha radiation. High labeling efficiencies and stability of 213Bi bound to human recombinant PAI-2 conjugated with cyclic diethylenetriaminepentaacetic acid anhydride were achieved (greater than 90%). The uPA inhibitory activity of the chelated PAI-2 was maintained as determined by complex formation with uPA and by inhibition of uPA activity. Furthermore, the reactivity of alpha-PAI-2 was confirmed in a cell assay as this construct was highly cytotoxic to breast cancer cell lines that express active, receptor bound uPA. The specificity of alpha-PAI-2 targeting was shown using several controls. Firstly, an active uPA blocking agent that limits PAI-2 binding significantly improved cell survival by a factor greater than three. Secondly, a non-specific alpha-BSA construct had minimal cytotoxic effect. Moreover, alpha-PAI-2 was not cytotoxic to freshly isolated normal human leukocytes, confirming that cells which do not contain active, receptor bound uPA cannot be targeted by alpha-PAI-2. In conclusion, we have validated, in vitro, the potential of alpha-PAI-2 as a novel therapeutic agent for breast cancer.
