The specific seroreactivity to ∆Np73 isoforms shows higher diagnostic ability in colorectal cancer patients than the canonical p73 protein

Abstract

The p53-family is tightly regulated at transcriptional level. Due to alternative splicing, up to 40 different theoretical proteoforms have been described for p73 and at least 20 and 10 for p53 and p63, respectively. However, only the canonical proteins have been evaluated as autoantibody targets in cancer patients for diagnosis. In this study, we have cloned and expressed in vitro the most upregulated proteoforms of p73, ΔNp73α and ΔNp73β, for the analysis of their seroreactivity by a developed luminescence based immunoassay test using 145 individual plasma from colorectal cancer, premalignant individuals and healthy controls. ∆Np73α seroreactivity showed the highest diagnostic ability to discriminate between groups. The combination of ∆Np73α, ∆Np73β and p73 proteoforms seroreactivity were able to improve their individual diagnostic ability. Competitive inhibition experiments further demonstrated the presence of unique specific epitopes in ΔNp73 isoforms not present in p73, with several colorectal patients showing unique and specific seroreactivity to the ΔNp73 proteoforms. Overall, we have increased the complexity of the humoral immune response to the p53-family in cancer patients, showing that the proteoforms derived from the alternative splicing of p73 possess a higher diagnostic ability than the canonical protein, which might be extensive for p53 and p63 proteins.

Identification of tumor-associated antigens with diagnostic ability of colorectal cancer by in-depth immunomic and seroproteomic analysis

“…This last limitation could produce the misidentification of relevant TAAs, which could be overcome by immunoprecipitation. Here, among the identified proteins as potential autoantibody targets in CRC (Table 2), we observed the presence of at least one protein containing continuous and discontinuous epitopes -p53-, as it has been previously demonstrated by ELISA, WB and protein microarrays [23,29,43], and thus supporting our approach to identify TAAs avoiding conformational limitations.…”

ΔNp73, TAp73 and Δ133p53 Extracellular Vesicle Cargo as Early Diagnosis Markers in Colorectal Cancer

“…We previously analyzed the potential seroreactivity of different variants of the p53 family (p53, p73, ΔNp73α, and ΔNp73β) [ 35 ] in a subset of our current cohort (healthy, n = 15; PL, n = 19; CRC, n = 9). Here, the correlation between the autoantibody levels and the ΔNp73 and Δ133p53 EV content was evaluated.…”

“…In our attempt to identify biomarkers for the early detection of CRC and the plausible usage of the TP53 family members to this aim, we have recently described a significant difference in the seroreactivity for ΔNp73 between CRC patients and subjects with PL versus healthy controls [ 35 ]. Interestingly, we have not observed a correlation between ΔNp73 seroreactivity and EVs’ mRNA levels of ΔNp73, TAp73 and Δ133p53, although a trend is observed for the negative association between the levels of ΔNp73 autoantibodies and EVs’ ΔNp73 mRNA in patients with CRC.…”

“…Among these samples, 15/29 of control samples, 19/49 of adenoma samples, and 9/42 of CRC samples were additionally used to study their humoral response in a previous work [ 35 ]. The clinical data of the subjects included in the study are shown in Table 1 .…”

Image-Based Network Analysis of DNp73 Expression by Immunohistochemistry in Rectal Cancer Patients

“…This negative regulation by DNp73 forms an autoregulatory feedback loop, since both TAp73 and p53 can induce expression of DNp73 isoforms by direct binding to the P2 promoter (Irwin, 2006;Rufini et al, 2011;Di et al, 2013). A newest evidence showed that DNp73 isoforms has higher applicant potential in colorectal cancer patients than the canonical p73 protein (Garranzo-Asensio et al, 2019). Thus, it is reasonable to focus on DNp73.…”