Generation of novel antibodies to delta p53 and evaluation in breast cancer cell lines and breast cancer specimens.

AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA 2787 Background: P53 is a tumor-suppressor gene that codes for a multi-functional DNA-binding protein whose primary role is to maintain genomic integrity through cell cycle arrest, DNA repair, and apoptosis. Approximately 20% of breast cancers contain a mutation within exons 5-8, which encompasses the DNA binding region of p53. Mutation in this region generally leads to aberrant folding and inactivity of the protein. Delta p53 however is an isoform that splices out part of exon 7 and all of exons 8 and 9, but results in a p53 isoform with altered transcriptional function. Delta p53 maintains the ATR-Intra-S phase checkpoint through transactivation of p21 and 14-3-3alpha, and this incurs the advantage of enhanced security in the DNA repair process by promoting coordinated repair and replication. Contrarily, if delta p53 occurs in conjunction with p53 mutations at aa273 and aa282 and DNA damage is present, then the function of p53 is abrogated, which may be associated with worse prognosis. For this reason we set out to generate antibodies that bind to delta p53. Results: Here we present the biochemistry for purifying native delta p53 protein and details on the RIMMS and contra-lateral immunization strategies to generate novel monoclonal antibodies. Immunoprecipitation studies indicate that the delta p53 monoclonal antibodies specifically pull down delta p53. In addition, we have demonstrated that these antibodies can be used to detect delta p53 in ELISA, IHC, and tumor tissue micorarrays assay formats. In a gamma irradiation tumor cell model, data indicates that all of the delta p53 antibodies react preferentially with delta p53. One antibody seems to be sensitive for a posttranslational modification occurring on both the wild type and delta p53 protein after irradiation. Conclusion: The presence of p53 mutations has been shown to affect prognosis of some breast cancers. Tools such as monoclonal antibodies are needed to identify delta p53 and these tools can be used to determine tumor clonality. Our findings indicate that our novel delta p53 antibodies may be useful in the evaluation and characterization of breast cancer tumors for response to radiation therapy or hormone-based therapy.