The Alpha-1-Fetoprotein (AFP) – A novel target gene for p63

Background Hepatocellular carcinoma (HCC) is the fifth common tumor entity worldwide. Despite advances in new technologies in both diagnosis and treatment, incidence and mortality continue to rise. So far, only orthotopic liver transplantation or surgical resection is curative. However, in many cases liver transplantation or resection of the tumor is not possible. Thus, novel treatment options are currently under investigation, e.g. combination treatments with checkpoint inhibitors and blockers of angiogenesis. Nevertheless, there is an urgent need for additional highly efficient approaches to treat HCC. Here, we analyze the role the Alpha-1-Fetoprotein (AFP) and its regulation by p53 family members to identify new possible therapeutic options to target HCC.

Methods In silico binding site analysis was done using the following databases: Transfac®, Jaspar. Hep3B cells were transfected with the following plasmids pcDNA-GFP (control), pcDNA TAp53, pcDNA TAp63, pcDNA TAp73 und pcDNA ∆Np73, respectively. mRNA was extracted using RNeasy Minikit (Qiagen). Upon reverse transcription cDNA was analyzed by quantitative PCR. Protein expression of p53 family members and AFP was analyzed by Western blot analysis.

Results Here, we show that AFP is a target protein of p53 family members. We identified in silico putative binding sites for p53, p63 and p73 in the promotor region and intron 1, 2, 3, and 4 of the AFP gene. Overexpression of p53, p73 and especially p63 lead to a downregulation of AFP expression on transcript (mRNA) level. In contrast, DNp73 has no influence on AFP expression. The data was verified by analysis of AFP protein expression. AFP protein levels were reduced after p53, p73 and p63 overexpression. Underlining the important role of p53 family members as negative regulators for AFP.

Conclusion Here, we demonstrate that p53 family members play an important role in regulation of AFP expression. Putative binding sites for all three family members (p53, p63 and p73) have been identified within the AFP gene. All full-length p53 family members are negative regulators for AFP expression. DNp73 did not show a regulatory effect on AFP expression. Of interest, p63 was identified as a novel negative regulator of AFP expression.

AFP is not only a tumor marker for HCC but also involved in downregulation of various immune responses. Thus, understanding the crosstalk of p53 family members and AFP expression shed new light on the pathophysiology of HCC.

Publication History

Article published online:
04 January 2021