S47 Variant Research Articles

Neither the African-Centric S47 Nor P72 Variant of TP53 Is Associated With Reduced Risk of Febrile Malaria in a Malian Cohort Study.

TP53 has been shown to play a role in inflammatory processes, including malaria. We previously found that p53 attenuates parasite-induced inflammation and predicts clinical protection to Plasmodium falciparum infection in Malian children. Here, we investigated whether p53 codon 47 and 72 polymorphisms are associated with differential risk of P. falciparum infection and uncomplicated malaria in a prospective cohort study of malaria immunity. p53 codon 47 and 72 polymorphisms were determined by sequencing TP53 exon 4 in 631 Malian children and adults enrolled in the Kalifabougou cohort study. The effects of these polymorphisms on the prospective risk of febrile malaria, incident parasitemia, and time to fever after incident parasitemia over 6 months of intense malaria transmission were assessed using Cox proportional hazards models. Confounders of malaria risk, including age and hemoglobin S or C, were similar between individuals with or without p53 S47 and R72 polymorphisms. Relative to their respective common variants, neither S47 nor R72 was associated with differences in prospective risk of febrile malaria, incident parasitemia, or febrile malaria after parasitemia. These findings indicate that p53 codon 47 and 72 polymorphisms are not associated with protection against incident P. falciparum parasitemia or uncomplicated febrile malaria.

Increased mTOR activity and metabolic efficiency in mouse and human cells containing the African-centric tumor-predisposing p53 variant Pro47Ser.

The Pro47Ser variant of p53 (S47) exists in African-descent populations and is associated with increased cancer risk in humans and mice. Due to impaired repression of the cystine importer Slc7a11, S47 cells show increased glutathione (GSH) accumulation compared to cells with wild -type p53. We show that mice containing the S47 variant display increased mTOR activity and oxidative metabolism, as well as larger size, improved metabolic efficiency, and signs of superior fitness. Mechanistically, we show that mTOR and its positive regulator Rheb display increased association in S47 cells; this is due to an altered redox state of GAPDH in S47 cells that inhibits its ability to bind and sequester Rheb. Compounds that decrease glutathione normalize GAPDH-Rheb complexes and mTOR activity in S47 cells. This study reveals a novel layer of regulation of mTOR by p53, and raises the possibility that this variant may have been selected for in early Africa.

Abstract 3483: The African-specific S47 variant of the p53 tumor suppressor gene alters cell metabolism

Abstract The TP53 gene, often referred to as the guardian of the genome, is the most frequently mutated gene in human cancer. This gene encodes the tumor suppressor p53, a master regulator of various processes such as programmed cell death, growth arrest and senescence. However, there is increasing evidence that highlights the role of p53 in tumor suppression with regards to maintaining metabolic homeostasis. Our lab has identified a polymorphic variant of p53 that encodes a serine residue instead of a proline at amino acid 47 (hereafter S47). This variant is most prevalent in African and African-American individuals, and mice and humans carrying this variant have increased cancer risk and impaired response to therapy. Using both mouse models and human cells, we have found that S47 cells have significantly altered metabolism compared to the WT counterpart. Specifically, S47 mice have increased weight and increased lean content compared to WT mice. These mice also show increased fitness in treadmill studies. Analyses of cell metabolism in S47 murine and human cells indicate that S47 cells show increased glycolysis and glycolytic flux, along with increased oxygen consumption rate on a Seahorse analyzer. The combined data support the premise that the S47 variant confers increased cancer risk but also increased metabolic fitness; the latter may explain the high rate of this variant in certain regions of Africa. By clearly elucidating the roles of WT p53 and the S47 variant in the context of metabolism, we hope to uncover new therapeutic avenues and ultimately enable more personalized medicine approaches for individuals who carry this variant. Citation Format: Keerthana Gnanapradeepan, Subhasree Basu, Che-Pei Kung, Thibaut Barnoud, Madeline Good, William Quinn, Joyce Lee, Kathryn Wellen, Zachary Schug, Joseph Baur, Donna George, Maureen Murphy. The African-specific S47 variant of the p53 tumor suppressor gene alters cell metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3483.

Elevated telomere dysfunction in cells containing the African-centric Pro47Ser cancer-risk variant of TP53.

Subtelomeric transcription and chromatin can have a significant impact on telomere repeat maintenance and chromosome stability. We have previously found that tumor suppressor protein p53 (TP53) can bind to retrotransposon-like elements in a majority of human subtelomeres to regulate TERRA transcription and telomeric histone acetylation in response to DNA damage. TP53 also prevents the accumulation of γH2AX DNA-damage signaling at telomeres. We now show that the inherited TP53 polymorphism Pro47Ser (hereafter S47), which is enriched in populations of African descent, is associated with elevated marks of telomere dysfunction. We found that human and mouse cells carrying the S47 variant show increased γH2AX DNA-damage signals at telomeres, as well as reduced TERRA transcription and subtelomeric histone acetylation in response to DNA damage stress. Cell-lines containing inducible genes for P47 or S47 versions of p53, as well mouse embryo fibroblasts (MEFs) reconstituted with human p53, showed elevated telomere-induced DNA damage foci and metaphase telomere signal loss in cells with S47. Human lymphoblastoid cell lines (LCLs) derived from individuals homozygous for S47, show increased accumulation of subtelomeric γH2AX and unstable telomere repeats in response to DNA damage relative to age matched LCLs homozygous for P47. Furthermore, LCLs with S47 had reduced replicative lifespan. These studies indicate that the naturally occurring S47 variant of p53 can affect telomeric chromatin, telomere repeat stability, and replicative capacity. We discuss the potential evolutionary significance of the S47 variant to African populations with respect to telomere regulation and the implications for inherited health disparities.

Mechanistic basis for impaired ferroptosis in cells expressing the African-centric S47 variant of p53

A population-restricted single-nucleotide coding region polymorphism (SNP) at codon 47 exists in the human TP53 gene (P47S, hereafter P47 and S47). In studies aimed at identifying functional differences between these variants, we found that the African-specific S47 variant associates with an impaired response to agents that induce the oxidative stress-dependent, nonapoptotic cell death process of ferroptosis. This phenotype is manifested as a greater resistance to glutamate-induced cytotoxicity in cultured cells as well as increased carbon tetrachloride-mediated liver damage in a mouse model. The differential ferroptotic responses associate with intracellular antioxidant differences between P47 and S47 cells, including elevated abundance of the low molecular weight thiols coenzyme A (CoA) and glutathione in S47 cells. Importantly, the disparate ferroptosis phenotypes related to the P47S polymorphism are reversible. Exogenous administration of CoA provides protection against ferroptosis in cultured mouse and human cells, as well as in a mouse model. The combined data support a positive role for p53 in ferroptosis and identify CoA as a regulator of this cell death process. Together, these findings provide mechanistic insight linking redox regulation of p53 to small molecule antioxidants and stress signaling pathways. They also identify potential therapeutic approaches to redox-related pathologies.

Tumor cells containing the African-Centric S47 variant of TP53 show increased Warburg metabolism.

Mutations in the TP53 tumor suppressor gene remain a hallmark of human cancer. In addition to mutation of TP53, single nucleotide polymorphisms (SNPs) in this gene can have a profound impact on p53 function, and can affect cancer risk as well as other p53 functions. Wild type (WT) p53 contains a proline at amino acid 47, but approximately 1% of African-Americans express a p53 allele with a serine at amino acid 47 (Pro47Ser, hereafter S47). In a mouse model for this variant, mice expressing S47 are predisposed to spontaneous cancers. The S47 variant also is associated with increased pre-menopausal breast cancer risk in African American women. We recently reported that S47 tumor cells are resistant to the majority of cytotoxic chemotherapeutic agents, but show increased sensitivity to a subset of anti-cancer agents, compared to tumors with WT p53. In this work, we report on another potentially promising therapeutic vulnerability of S47 tumors. We find that S47 tumors show decreased mitochondrial metabolism, along with increased dependency on glycolysis. S47 tumor cells also show increased sensitivity to the glycolytic poison 2-deoxy-glucose. We propose that the altered metabolism in S47 tumor cells may be yet another potentially-actionable therapeutic vulnerability to exploit in cancer-prone individuals with this genotype.

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.

The transcription-independent mitochondrial cell death pathway is defective in non-transformed cells containing the Pro47Ser variant of p53

ABSTRACTApproximately half of all human cancers contain mutations in the TP53 tumor suppressor. In addition to mutations, there are single nucleotide polymorphisms (SNPs) in TP53 that can dampen p53 function, and can increase cancer risk and decrease the efficacy of cancer therapy. Approximately 6% of Africans and 1% of African-Americans express a p53 allele with a serine instead of proline at position 47 (Pro47Ser, or S47). The S47 variant is associated with increased breast cancer risk in pre-menopausal African Americans, and in a mouse model for the S47 variant, mice are predisposed to spontaneous cancers. We recently showed that the S47 variant is impaired for p53-mediated apoptosis in response to radiation and some genotoxic agents, particularly cisplatin. Here we identify the mechanism for impaired apoptosis of S47 in response to cisplatin. We show that following cisplatin treatment, the S47 variant shows normal stabilization and serine 15 phosphorylation, but reduced ability to bind to the peptidyl prolyl isomerase PIN1, which controls the mitochondrial localization of p53. This is accompanied by impaired mitochondrial localization of S47, along with decreased induction of cleaved caspase-3. Interestingly, we show that this defect occurs only for cisplatin and not for camptothecin. These findings show that normal tissues may respond differently to genotoxic stress depending upon this TP53 genotype. These data suggest that toxicity to cisplatin may be decreased in S47 individuals, and that this compound may be a superior treatment option for these individuals.

Abstract B210: BET inhibitors show superior ability to suppress the progression of tumors containing the Pro47Ser single-nucleotide polymorphism of p53

Abstract The tumor suppressor TP53 is the most frequently mutated gene in human cancer and serves to restrict both tumor initiation and progression. Single-nucleotide polymorphisms (SNPs) in the p53 pathway can have a marked impact on p53 tumor suppressor function, and several have been associated with increased cancer risk. We have reported that a naturally occurring SNP affects codon 47 of TP53 (rs1800371, G/A). In most individuals, codon 47 encodes a proline (P47). However, approximately 2.5% of African-Americans express a p53 allele with a serine residue (S47) at this position. In order to compare the ability of the WT and S47 variants of p53 to suppress tumor development, we isolated mouse embryonic fibroblasts (MEFs) from WT and S47 mice and transformed them with the oncogenes E1A and Ras, in order to generate a facile model that would allow us to study responses to therapy. WT and S47 cells grew similarly in 10% serum. However, transformed S47 cells grew significantly better under nutrient-deprived conditions. S47 transformed cells also grew with increased dynamics in a xenograft mouse model and in a tail vein model of metastasis. These data support the premise that the S47 variant of p53 is significantly impaired as a tumor suppressor. We found that, for the majority of conventional genotoxic drugs, S47 tumor cells are resistant, compared to tumor cells with WT p53. In order to identify novel compounds that preferentially target S47 tumor cells, we performed a drug screen on WT and S47 transformed MEFs, using a library of over 200 conventional and experimental chemotherapeutic drugs. We found that S47 cells are far more sensitive to the class of bromodomain inhibitors, specifically BET inhibitors. Using short-term (96 hrs, Alamar Blue) cell viability assays, we found that S47 transformed cells were approximately 5-fold more sensitive to OTX-015, a BET inhibitor currently in clinical trials. We also found that S47 cells were significantly impaired in their ability to form colonies when pretreated for 24 hours with OTX-015. These data highlight the relevance of targeted therapy for patients harboring the S47 variant of p53, and suggest that BET inhibitors may preferentially target S47 tumors. Citation Format: Thibaut Barnoud, Subhasree Basu, Che-Pei Kung, Matthew Reiss, Rugang Zhang, Maureen Murphy. BET inhibitors show superior ability to suppress the progression of tumors containing the Pro47Ser single-nucleotide polymorphism of p53 [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B210.

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.

Abstract 2964: The codon 47 Pro47Ser polymorphism influences p53 phosphorylation and apoptotic function

Abstract The p53 tumor suppressor protein is a central signaling protein in response to oncogenic, genotoxic and metabolic stress. Single nucleotide polymorphisms (SNPs) in p53 can have a marked impact on p53 tumor suppressor function, and have been associated with increased cancer risk. Amino acid 47 of p53 is a conserved residue, located within the second transactivation domain of this protein. In wild type p53, this residue is proline (P47, or WT). Importantly, this proline is critical for the ability of proline-directed kinases to phosphorylate p53 on the adjacent residue, serine 46. Phosphorylation of serine 46 of p53 greatly enhances the ability of p53 to transactivate certain target genes, and to induce apoptosis. In some Africans and African Americans, amino acid 47 of p53 encodes serine (S47) instead of proline. We have shown that the S47 polymorphism eliminates phosphorylation on serine 46, and reduces p53's ability to induce apoptosis by at least 3-fold. We have created mice that contain the humanized p53 gene (Hupki) for either the S47 or WT alleles of p53. In new data, we show that tissues from these mice have significantly decreased apoptosis after gamma radiation. We also show that apoptosis induced by cisplatin is markedly defective in S47 mice and cell lines. Further, MEFs from S47 mice have a 15-fold increase in IC50 for cisplatin, indicating that this polymorphism will impact the efficacy of cisplatin therapy in Africans and African Americans. Finally, we have identified two critical p53 target genes, GLS2 (metabolism) and NOXA (apoptosis), which show significantly impaired transactivation by S47. The long-term objective of this research is to further characterize the function of the S47 variant, and to identify the contribution of this variant to cancer risk and progression in Africans and African Americans. Citation Format: Matthew Jennis, Monica Hollstein, Maureen E. Murphy. The codon 47 Pro47Ser polymorphism influences p53 phosphorylation and apoptotic function. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2964. doi:10.1158/1538-7445.AM2014-2964