p53 and cancer

3.5.1 p53 mutant and tumor formation

Inactivation of the p53 tumor suppressor is a frequent event in tumorigenesis. As reported, p53 mutations are found in 50–55% of all human cancers, demonstrating the crucial role of p53 in tumor suppression (Hamzehloie et al., 2012) (for further information, see the IARC TP53 mutation database (http://p53.iarc.fr/TP53SomaticMutations.aspx). Unlike the majority of tumor suppressor genes, such as RB, APC, or BRCA1, which are usually inactivated during cancer progression by deletions or truncating mutations, the TP53 gene in human tumors is often found to undergo missense mutations, in which a single nucleotide is substituted by another. Consequently, a full-length protein containing only a single amino acid substitution is produced (Hainaut and Hollstein, 2000).

The cancer-associated TP53 mutations are very diverse in their locations within the p53 coding sequence and their effects on the thermodynamic stability of the p53 protein. However, the vast majority of the mutations result in loss of p53 ability to bind DNA in a sequence-specific manner and activate transcription of canonical p53 target genes (Bullock and Fersht, 2001). TP53 mutations are distributed in all coding exons, with a strong predominance in exons 4-9, which encode the DNA-binding domain of the protein. Of the mutations in this domain, about 30% fall within 6 “hotspot” residues (residues R175, G245, R248, R249, R273, and R282) and are frequent in almost all types of cancer (Cho et al., 1994). The existence of these hotspot residues could be explained both by the susceptibility of particular codons to carcinogen-induced alterations and by positive selection of mutations that affect cell growth, thus providing survival advantages. In addition to the loss of function that a mutation in TP53 may cause, many p53 mutants are able to actively promote tumor development by several other means. In a heterozygous situation, where both WT and mutant alleles exist, mutant p53 can antagonize WT p53 tumor suppressor functions in a dominant negative (DN) manner. The

fact that the transcriptional activity of WT p53 relies on the formation of tetramers, whose DNA binding function may be interfered by mutant p53 (Milner et al., 1991;

Sigal and Rotter, 2000). However, such a heterozygous state is often transient, as TP53 mutations are frequently followed by loss of heterozygosity (LOH) during cancer progression. LOH is often seen in the case of tumor suppressors where, at a particular locus heterozygous for a mutant and WT allele, the WT allele is either deleted or mutated. The LOH of the short arm of chromosome 17, where TP53 is located (Miller et al., 1986), implies a selective force driving the inactivation of the remaining WT allele, suggesting that the DN activity of mutant p53 is not sufficient to completely inactivate WT p53. Furthermore, data supports the concept that many mutant p53 isoforms can exert additional oncogenic activity by a gain-of-function (GOF) mechanism. GOF refers to the acquisition of oncogenic properties by the mutant protein, compared with the mere inactivation of the protein (Brosh and Rotter, 2009; Oren and Rotter, 2010). Both the DN and GOF effects may play a significant role in the positive selection of missense mutations in TP53 during tumorigenesis (Miller et al., 2016). Mutant p53 oncogenic properties of and their underlying mechanisms are shown in Figure 30.

Figure 30. Selected oncogenic properties of mutant p53 and their underlying mechanisms. The inner circle (shaded blue) represents oncogenic phenotypes associated with the activities of mutant p53 proteins. The outer circle depicts key mechanistic properties of p53 mutants that underlie the phenotypes listed in the inner circle. Note that each of the phenotypic effects can be attributed to almost each of the mechanistic properties; hence the inner blue circle can be freely rotated (Brosh and Rotter, 2009).

Inherited TP53 mutations are associated with a rare autosomal dominant disorder, the Li-Fraumeni syndrome (LFS). LFS is unusual because it causes susceptibility to a wide variety of cancers. It is characterized by multiple primary neoplasms in children and young adults, with a predominance of soft-tissue sarcomas, osteosarcomas, breast cancers, brain tumors and adrenocortical carcinomas (Li et al., 1988; Valdez et al., 2017). In about 70% of these multicancer families, mutant alleles of p53 were found to be transmitted in a mendelian fashion. Family members who inherited a mutant p53 allele had a high probability of developing some form of malignancy, often early in life. The age of onset of these various malignancies was found to be quite variable: about 5 years of age for adrenocortical

carcinomas, 16 years for sarcomas, 25 years for brain tumors, 37 years for breast cancer, and almost 50 years for lung cancer (Weinberg, 2013).

3.5.2 p53 and cancer treatment

The TP53 gene is considered as one of the most important and well-known tumour suppressor gene and is the most frequently inactivated protein in human cancers, as it is mutated in 50% of human tumours (Leroy et al., 2014). Despite the high mutation rate of the protein p53, the remaining half of human tumours retains wild-type p53, implying that these tumours might have developed alternative mechanisms for disabling or attenuating specific p53 functions or the entire p53 pathway. This is also possible because p53 activity is strictly regulated, to avoid apoptosis induction in normal growth conditions. One of these regulatory mechanisms is represented by the ubiquitin-dependent degradation. Indeed, under non-stressed conditions, p53 is tightly regulated by the E3 ubiquitin ligase MDM2 protein through an auto-regulatory feedback loop, as this protein is also a p53 transcriptional target (Chene, 2003; Freedman et al., 1999; Picksley and Lane, 1993). Binding of MDM2 to the transactivation domain of p53 promotes the nuclear export of p53 and serves as a ubiquitin ligase that promotes ubiquitin-dependent degradation of p53 (Mendoza et al., 2014). The MDM2 gene has been found amplified or overexpressed in 7% of human malignancies, particularly in soft tissue tumours, testicular germ cell cancers and neuroblastoma (Oliner et al., 2016). It acts as an oncogene altering the p53 pathway especially in wild-type p53 tumour cells. It could then explain the resistance to cancer cell death in tumour cells expressing wild-type p53. Activation of p53 in human cancers harbouring wild-type p53 may offer a therapeutic benefit to induce cancer cell death. For this purpose, several small molecules able to inhibit p53-MDM2 binding have been developed, such as Nutlins, which activate the p53 pathway in vitro and in vivo (Vassilev et al., 2004). However, anti-tumour efficiency of p53 activation requires not only the presence of wild-type p53, but also functional p53 signalling pathway.

Importantly, some other mechanisms have been developed in cancer cells to block apoptosis induced by p53, as the case for the couple Netrin-1/UNC5B, data in our lab strongly showing that an increase of Netrin-1 gene expression and a resistance to apoptosis upon p53 activation and induction (Paradisi et al., 2013). In addition, some papers also report that target to p53-dependent receptors-ligand complex, like UNC5B-Netrin-1 can also induce apoptosis dependent on the balance between the expression of receptor and ligand (Tanikawa et al., 2003).