Role of p73 Dinucleotide Polymorphism in Prostate Cancer and p73 Protein Isoform Balance

Abstract

Background. Molecular markers for prostate cancer (PCa) risks are currently lacking. Here we address the potential association of a dinucleotide polymorphism (DNP) in exon 2 of the p73 gene with PCa risk/progression and discern any disruption of p73 protein isoforms levels in cells harboring a p73 DNP allele. Methods. We investigated the association between p73 DNP genotype and PCa risk/aggressiveness and survival by fitting logistic regression models in 1,292 incident cases and 682 controls. Results. Although we detected no association between p73 DNP and PCa risk, a significant inverse relationship between p73 DNP and PCa aggressiveness (AT/AT + GC/AT versus GC/GC, OR = 0.55, 95%Cl = 0.31-0.99) was detected. Also, p73 DNP is marginally associated with overall death (dominant model, HR = 0.76, 95%Cl = 0.57-1.00, P = 0.053) as well as PCa specific death (HR = 0.69, 95%Cl = 0.45-1.06, P = 0.09). Western blot analyses for p73 protein isoforms indicate that cells heterozygous for the p73 DNP have lower levels of ∆Np73 relative to TAp73 (P < 0.001). Conclusions. Our findings are consistent with an association between p73 DNP and low risk for PCa aggressiveness by increasing the expressed TAp73/∆Np73 protein isoform ratio.

Figure 1

Association between rs1801173 and overall survival (a) and prostate cancer specific survival (b). A role of p73 DNP in prostate cancer progression and aggressiveness was investigated. p73 DNP is marginally associated with overall death (dominant model, hazard ratio (HR) = 0.76, 95%CI = 0.57–1.00, P = 0.053 by Cox regression adjusted for age) as well as PCa specific death (HR = 0.69, 95%CI = 0.45–1.06, P = 0.09 by the competing risk regression adjusted for age).

Figure 2

Western analyses of p73 protein N-terminal isoforms in cancer cell lines. Proteins from cancer cell line lysates in Figure 2(a) (1.25 μg/well) and Figure 2(b) (7.50 μg/well) were resolved on 10% SDS-PAGE gels and the resolved proteins were electrotransferred onto PVDF membranes and then immunoblotted (IB) with a p73 isoform-specific monoclonal antibody against TAp73 or ΔNp73. As a positive loading control, actin was immunoblotted with a goat polyclonal antibody. Primary antibodies were detected using appropriate secondary antibody-horseradish peroxidase conjugates and an enhanced chemoluminescence method followed by fluorography (IB = primary western antibody).

Figure 3

Relative TAp73/ΔNp73 protein isoform ratios in cancer cell line lysates with GC/GC and GC/AT genotype. The TAp73/ΔNp73 protein isoform levels were determined from the western blotting data in Figures 2(a) and 2(b). These p73 protein isoform levels were used to calculate the relative ratio values reported in Table 3. The mean of TAp73/ΔNp73 protein isoform ratio values is box-plotted.