ABSTRACT
The tumor suppressor gene p53 is mutated in more than half of human tumors. One important characteristic of p53 mutants is their accumulation in the nucleus of cancer cells. Thus, reactivation of mutant p53 proteins may trigger massive apoptosis in tumor cells. Pharmacologic methods are currently under development to induce mutant p53 proteins to resume their wild-type function. We have identified a human single-chain Fv fragment, designated as transcriptional transactivation and apoptosis restoring (TAR1), which specifically and with high affinity binds to mutant p53 and restores its wild-type active conformation. Binding of TAR1 to mutant p53 induced transcriptional transactivation of p53 target genes and down-regulation of mutant p53 transcriptional target genes. TAR1 treatment induced apoptosis in a variety of cell lines endogenously expressing p53 carrying different point mutations DNA contact or structural p53 mutants. Moreover, in an animal model of mice carrying human xenografts, TAR1 induced tumor regression with no apparent deleterious side effects. Thus, it may be considered as a potential candidate for anticancer treatment, targeting tumors with mutant p53.
Subject(s)
Mutant Proteins/immunology , Mutant Proteins/metabolism , Neoplasms, Experimental/immunology , Single-Chain Antibodies/administration & dosage , Tumor Suppressor Protein p53/immunology , Animals , Apoptosis/drug effects , Caspase 3/immunology , Caspase 3/metabolism , Caspase 7/immunology , Caspase 7/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Humans , Mice , Mice, Nude , Mutant Proteins/genetics , Neoplasms, Experimental/genetics , Neoplasms, Experimental/metabolism , Neoplasms, Experimental/pathology , Protein Binding , Signal Transduction/drug effects , Single-Chain Antibodies/chemistry , Single-Chain Antibodies/isolation & purification , Single-Chain Antibodies/pharmacology , Transcriptional Activation , Tumor Burden/drug effects , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , Xenograft Model Antitumor AssaysABSTRACT
Here, we describe the biological activity of ME1, a mouse single chain Fv fragment (scFv) against the common epitope of mutant p53, which is efficiently expressed in mammalian cells. We found that in vivo interaction of the conformational p53 mutant R175H protein with the scFv resulted in the acquisition of wild-type p53 characteristics, manifested in trans-activation of p21, as well as induction of apoptosis. Moreover, antibody binding leads to abrogation of the mutant p53 mediated "gain of function" as estimated by downregulation of EGR-1, a transcriptional target of mutant p53. These findings suggest that the scFv restores wild-type properties to mutant p53.
