Abstract 881: Identification of autoantibody biomarkers to wild-type and mutant p53 in pancreatic and ovarian cancer

Abstract

Abstract Introduction: Mutations in TP53 are common in early pancreatic and ovarian cancer development, and are detected in the majority of pancreatic cancers (75%) and serous ovarian cancers (97%). Mutations in TP53 induce autoantibody (AAb) immune responses, which may be biomarkers for the early detection of these cancers. The most common mutations in p53 occur in the DNA binding domain at residues that directly contact the DNA (Arg175, 248, and 273). In contrast, preliminary studies suggest that mutations in the N- and C- termini of p53 alter the thermodynamic stability of p53, leading to increased accumulation in tumors. It is not known if the location of the mutations impacts immunogenicity, or if p53-AAb are specific to mutant p53 proteins. Methods: To evaluate AAb to mutant p53 proteins, we generated custom programmable protein microarrays displaying 52 common p53 point mutations. cDNAs encoding full-length p53 with c-terminal GST tags were printed on arrays with anti-GST antibody, and transcribed and translated in situ using HeLa expression lysate. The microarrays were screened with sera from pancreatic cancer cases (Stage I-III, n=60), healthy and benign disease controls (n=63), ovarian cancer cases (n=19) and controls (n=17). Bound IgG was detected by fluorescence. For validation, four immunogenic mutant p53 proteins Arg248Met, Pro278Ser, Arg213Leu, Gly187Ser and wild-type p53 were individually displayed in 96-well format using a programmable ELISA. The ELISAs were re-screened with a subset of the sera from pancreatic cancer cases (n=49) and controls (n=47) and all of the ovarian cancer cases and controls. Cutoff values were established to determine if a patient is positive for all wild-type and mutant p53 AAb by measuring the mean antigen florescence intensity +2 standard deviations of the controls. Results: AAb were detected to all four mutant and wild-type p53 proteins in sera from 8/49 (16%) of pancreatic cases and 2/47 (4.2%) of pancreatic controls, but the results were not statistically significant. In comparison, AAb to all four mutant and wild-type p53 proteins were identified in sera from 8/19 (42%) of ovarian cases and 0/17 (0%) of healthy control women (p ≤ 0.005). All sera with AAb to mutant p53 also had measurable AAb to wild-type p53 protein. All patients with immune responses had evidence of broad-reactivity to at least at least two mutant p53 proteins, as well as to wild-type p53. Conclusion: Our results indicate that patients with serum AAb to wild type p53 protein also have AAb to multiple mutant p53 antigens, suggesting degeneracy of the epitope specificity. No patients have been identified with AAb specific to mutant-only p53. We observed inter-patient variation in the patterns of immune reactivity to p53 mutations that is of unclear significance. Correlation with tumor p53 mutation status is ongoing. Citation Format: Benjamin A. Katchman, Rizwan Alam, Garrick Wallstrom, Joshua LaBaer, Michael A. Hollingsworth, Daniel W. Cramer, Karen S. Anderson. Identification of autoantibody biomarkers to wild-type and mutant p53 in pancreatic and ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 881. doi:10.1158/1538-7445.AM2014-881