Subhasree Basu

An African-specific polymorphism in the TP53 gene impairs p53 tumor suppressor function in a mouse model

A nonsynonymous single-nucleotide polymorphism at codon 47 in TP53 exists in African-descent populations (P47S, rs1800371; referred to here as S47). Here we report that, in human cell lines and a mouse model, the S47 variant exhibits a modest decrease in apoptosis in response to most genotoxic stresses compared with wild-type p53 but exhibits a significant defect in cell death induced by cisplatin. We show that, compared with wild-type p53, S47 has nearly indistinguishable transcriptional function but shows impaired ability to transactivate a subset of p53 target genes, including two involved in metabolism:Gls2(glutaminase 2) and Sco2 We also show that human and mouse cells expressing the S47 variant are markedly resistant to cell death by agents that induce ferroptosis (iron-mediated nonapoptotic cell death). We show that mice expressing S47 in homozygous or heterozygous form are susceptible to spontaneous cancers of diverse histological types. Our data suggest that the S47 variant may contribute to increased cancer risk in individuals of African descent, and our findings highlight the need to assess the contribution of this variant to cancer risk in these populations. These data also confirm the potential relevance of metabolism and ferroptosis to tumor suppression by p53.

The P72R Polymorphism of p53 Predisposes to Obesity and Metabolic Dysfunction

p53 is well known for its tumor suppressor role, but this protein also has a poorly understood role in the regulation of metabolism. Human studies have implicated a common polymorphism at codon 72 of p53 in diabetic and pre-diabetic phenotypes. To understand this role, we utilized a humanized mouse model of the p53 codon 72 variants and monitored these mice following challenge with a high-fat diet (HFD). Mice with the arginine 72 (R72) variant of p53 developed more-severe obesity and glucose intolerance on a HFD, compared to mice with the proline 72 variant (P72). R72 mice developed insulin resistance, islet hypertrophy, increased infiltration of immune cells, and fatty liver disease. Gene expression analyses and studies with small-molecule inhibitors indicate that the p53 target genes Tnf and Npc1l1 underlie this phenotype. These results shed light on the role of p53 in obesity, metabolism, and inflammation.

Genetic Modifiers of the p53 Pathway

The tumor suppressor gene TP53 is the most frequently mutated gene in human cancer; this gene is subject to inactivation by mutation or deletion in >50% of sporadic cancers. Genes that encode proteins that regulate p53 function, such as MDM2, MDM4, and CDKN2A (p14(ARF)) are also frequently altered in tumors, and it is generally believed that the p53 pathway is likely to be inactivated by mutation in close to 100% of human tumors. Unlike most other cancer-relevant signaling pathways, some of the genes in the p53 pathway contain functionally significant single nucleotide polymorphisms (SNPs) that alter the amplitude of signaling by this protein. These variants, thus, have the potential to impact cancer risk, progression, and the efficacy of radiation and chemotherapy. In addition, the p53 pathway plays a role in other biological processes, including metabolism and reproductive fitness, so these variants have the potential to modify other diseases as well. Here we have chosen five polymorphisms in three genes in the p53 pathway for review, two in TP53, two in MDM2, and one in MDM4. These five variants were selected based on the quality and reproducibility of functional data associated with them, as well as the convincingness of epidemiological data in support of their association with disease. We also highlight two other polymorphisms that may affect p53 signaling, but for which functional or association data are still forthcoming (KITLG and ANRIL). Finally, we touch on three questions regarding genetic modifiers of the p53 pathway: Why did these variants arise? Were they under selection pressure? And, is there compelling evidence to support genotyping these variants to better predict disease risk and prognosis?

p53 family members regulate cancer stem cells

Cancer stem cells are commonly defined as those cells that constitute a very low fraction of cells in a tumor, and which are uniquely capable of generating a new tumor following transfer or migration into a new environment. For certain cancers such as colorectal, breast and certain leukemias, the existence of cancer stem cells is well-documented. Such cells are believed to be largely responsible for metastases, and for relapse following chemotherapy. The recent findings by some groups that some commonly-used chemotherapeutic agents can actually enrich for cancer stem cells has prompted the emerging concept that in order to successfully combat a tumor, both the bulk tumor and the cancer stem cells need to be targeted. The difficulty lies in finding the target: a protein expressed in both the bulk tumor and the stem cells, on which both the tumor and the cancer stem cell rely for survival. Two recent publications have identified an isoform of the p53 family member p73 as such a protein (Fig. 1). p53 has long been established as a negative regulator of stem cells. Additionally, p53 has 2 ‘siblings’, p63 and p73, which are homologous in structure and function. However, p63 and p73 have “Jekyll and Hyde” isoforms when it comes to tumor growth: namely, the full length isoforms, so-called TA p63 and TA p73, are predominantly tumor suppressive, while the N-terminal truncation variants, so-called DN-p63 and DN-p73, are typically tumor-promoting, and not surprisingly are frequently overexpressed in tumors. Two groups recently discovered that silencing DN-p73 causes decreased expression of stem cell markers, decreased anchorage-independent growth (a hallmark of cancer stem cell function) and decreased ability to engraft as tumors. In the publication by P€ utzer and colleagues, these researchers found that overexpression of DN-p73 in the melanoma Sk-Mel-29 led to higher CD133, Nanog and Oct4, along with increased anchorage independent growth and ability to form tumors in xenografts. Mechanistically this group found that the ability of DN-p73 to confer cancer stem cell function relied on its ability to negatively regulate the microRNA miR885-5p. They went on to show that the target of this microRNA is the IGF1 receptor. In sum, overexpression of DN-p73 variant led to greatly enhanced IGF1 signaling, which then functioned in a feedback loop to increase cancer stem cell function. El-Deiry and colleagues found a similar impact of DN-p73 on cancer stem cells, but this time focused on colorectal cancer stem cells. This group started with an innovative screen to search for compounds that could restore p53 function. These researchers discovered a compound called prodigiosin, which turned out to restore p53 function by inhibiting its negative regulator, DNp73. Mechanistically, this group found that the ability of prodigiosin to inhibit the function of DN-p73 occurred by virtue of the ability of this compound to increase the level of c-jun; the latter transcription factor upregulates TA-p73, which in turn inhibits DN-p73. El-Deiry and colleagues found that treatment of colorectal tumors with prodigiosin led to decreased colonosphere formation, decreased levels of cells positive for the stem cell markers ALDH, CD44, ID3 and E-cadherin, and decreased ability of colorectal cancer stem cells to initiate tumors as xenografts. Notably, when treatment with prodigiosin was stopped, tumor growth did not renew, supporting the premise that prodigiosin led to a cure in treated mice. These findings suggest that prodigiosin possesses unique anti-cancer functions.

The African-specific S47 polymorphism of p53 alters chemosensitivity

ABSTRACT The TP53 protein is known to affect the sensitivity of tumor cells to cell death by DNA damaging agents. We recently reported that human and mouse cells containing an African-specific coding region variant of p53, Pro47Ser (hereafter S47), are impaired in the transactivation of a small subset of p53 target genes including GLS2 and SCO2, and are markedly resistant to cisplatin. Further, mice containing this variant are markedly predisposed to cancer. Together these findings suggested that cancer-affected humans with the S47 variant might not be effectively treated with cisplatin. To more directly test this premise, we created transformed derivatives of mouse embryo fibroblasts (MEFs) containing wild type p53 (WT) and the S47 variant and analyzed them for chemosensitivity. We find that transformation with E1A and Ras actually reverses the chemosensitivity/transcriptional differences between WT p53 and S47. Specifically, E1A/Ras-transformed S47 cells show increased sensitivity to cisplatin and paclitaxel, and comparable transactivation of GLS2 and SCO2, compared to cells with WT p53. These data suggest that the functional differences between WT p53 and S47 in primary cells may not hold true for transformed cells. They also offer hope that cisplatin and paclitaxel may be effective chemotherapeutic drugs for S47 individuals with cancer.

The p53 Tumor Suppressor in the Control of Metabolism and Ferroptosis

The p53 tumor suppressor continues to be distinguished as the most frequently mutated gene in human cancer. It is widely believed that the ability of p53 to induce senescence and programmed cell death underlies the tumor suppressor functions of p53. However, p53 has a number of other functions that recent data strongly implicate in tumor suppression, particularly with regard to the control of metabolism and ferroptosis (iron- and lipid-peroxide-mediated cell death) by p53. As reviewed here, the roles of p53 in the control of metabolism and ferroptosis are complex. Wild-type (WT) p53 negatively regulates lipid synthesis and glycolysis in normal and tumor cells, and positively regulates oxidative phosphorylation and lipid catabolism. Mutant p53 in tumor cells does the converse, positively regulating lipid synthesis and glycolysis. The role of p53 in ferroptosis is even more complex: in normal tissues, WT p53 appears to positively regulate ferroptosis, and this pathway appears to play a role in the ability of basal, unstressed p53 to suppress tumor initiation and development. In tumors, other regulators of ferroptosis supersede p53’s role, and WT p53 appears to play a limited role; instead, mutant p53 sensitizes tumor cells to ferroptosis. By clearly elucidating the roles of WT and mutant p53 in metabolism and ferroptosis, and establishing these roles in tumor suppression, emerging research promises to yield new therapeutic avenues for cancer and metabolic diseases.

A link between TP53 polymorphisms and metabolism

ABSTRACT Besides being a critical tumor suppressor, the TP53 gene also plays a role in metabolism and recent studies in humans have implicated the codon 72 polymorphism of TP53 in this role. Using a humanized knock-in mouse model for these TP53 variants, we show that this polymorphism has a significant impact on the metabolic response to a high-fat diet.

PUMA-dependent apoptosis in NSCLC cancer cells by a dimeric β-carboline

Dimeric β-carbolines are cytotoxic against multiple NSCLC cell lines, and we report herein our preliminary studies on the mechanism of action of these dimeric structures. Dimeric β-carboline 1, which is more potent than the corresponding monomer in NSCLC cell lines, is a lysosomotropic agent that inhibits autophagy and mediates cell death by apoptosis, upregulating the pro-apoptotic BH3-only protein PUMA (p53 upregulated modulator of apoptosis) in a dose dependent manner.

The Hsp70 Family of Heat Shock Proteins in Tumorigenesis: From Molecular Mechanisms to Therapeutic Opportunities

The HSP70 family of molecular chaperones consists of at least eight members that are highly evolutionarily conserved. Whereas more than one member of this family is implicated in cancer, the most compelling and abundant data point to the involvement of the predominant stress-inducible form of this protein in cancer etiology and progression. High levels of HSP70 staining in tumors emerged as a significant marker of poor prognosis in human tumors over 20 years ago. Since that time, the important role of this protein in cellular transformation, viral infection, immune function, and the cellular stress response has come to be appreciated and understood. In the past 10 years, the findings that many different types of human tumors are addicted to this protein for survival, and that silencing HSP70 is cytotoxic to tumor but not normal cells, have led to the emergence of the first specific inhibitors of this family of molecular chaperones for cancer therapy. Here-in we review the pro-tumorigenic function(s) of this protein, our understanding of how HSP70 mediates protein quality control, and the current efforts to target and inhibit this protein for cancer therapy.

Tailoring Chemotherapy for the African-Centric S47 Variant of TP53

The tumor suppressor TP53 is the most frequently mutated gene in human cancer and serves to restrict tumor initiation and progression. Single-nucleotide polymorphisms (SNP) in TP53 and p53 pathway genes can have a marked impact on p53 tumor suppressor function, and some have been associated with increased cancer risk and impaired response to therapy. Approximately 6% of Africans and 1% of African Americans express a p53 allele with a serine instead of proline at position 47 (Pro47Ser). This SNP impairs p53-mediated apoptosis in response to radiation and genotoxic agents and is associated with increased cancer risk in humans and in a mouse model. In this study, we compared the ability of wild-type (WT) and S47 p53 to suppress tumor development and respond to therapy. Our goal was to find therapeutic compounds that are more, not less, efficacious in S47 tumors. We identified the superior efficacy of two agents, cisplatin and BET inhibitors, on S47 tumors compared with WT. Cisplatin caused dramatic decreases in the progression of S47 tumors by activating the p53/PIN1 axis to drive the mitochondrial cell death program. These findings serve as important proof of principle that chemotherapy can be tailored to p53 genotype.Significance: A rare African-derived radioresistant p53 SNP provides proof of principle that chemotherapy can be tailored to TP53 genotype. Cancer Res; 78(19); 5694-705. ©2018 AACR.