scholarly journals Sex- and Mutation-Specific p53 Gain-of-Function Activity in Gliomagenesis

2021 ◽  
Vol 1 (3) ◽  
pp. 148-163
Author(s):  
Nathan C. Rockwell ◽  
Wei Yang ◽  
Nicole M. Warrington ◽  
Max V. Staller ◽  
Malachi Griffith ◽  
...  
Keyword(s):  
Sex Differences ◽  
Dna Binding ◽  
Sex Difference ◽  
P53 Mutation ◽  
Mutant P53 ◽  
Missense Mutations ◽  
P53 Mutations ◽  
Primary Mouse

In cancer, missense mutations in the DNA-binding domain of TP53 are common. They abrogate canonical p53 activity and frequently confer gain-of-oncogenic function (GOF) through localization of transcriptionally active mutant p53 to noncanonical genes. We found that several recurring p53 mutations exhibit a sex difference in frequency in patients with glioblastoma (GBM). In vitro and in vivo analysis of three mutations, p53R172H, p53Y202C, and p53Y217C, revealed unique interactions between cellular sex and p53 GOF mutations that determined each mutation's ability to transform male versus female primary mouse astrocytes. These phenotypic differences were correlated with sex- and p53 mutation–specific patterns of genomic localization to the transcriptional start sites of upregulated genes belonging to core cancer pathways. The promoter regions of these genes exhibited a sex difference in enrichment for different transcription factor DNA-binding motifs. Together, our data establish a novel mechanism for sex-specific mutant p53 GOF activity in GBM with implications for all cancer. Significance: Sex differences in cancer, including glioblastoma, have been observed in both incidence and outcome. We reveal that TP53, the most commonly mutated gene in cancer, contributes to sex differences through differential GOF activity. This discovery has critical implications for our understanding of p53 mutations and the importance of sex as a biological variable.

scholarly journals p53 mutations exhibit sex specific gain-of-function activity in gliomagenesis

2021 ◽  
Author(s):  
Nathan C Rockwell ◽  
Wei Yang ◽  
Nicole Warrington ◽  
Malachi Griffith ◽  
Obi L Griffith ◽  
...  
Keyword(s):  
Dna Binding ◽  
Expression Patterns ◽  
Metabolic Reprogramming ◽  
Mutant P53 ◽  
Small Male ◽  
Missense Mutations ◽  
P53 Mutations ◽  
Gain Of Function ◽  
Primary Mouse

The tumor suppressor TP53 is the most frequently mutated gene in cancer. Most TP53 mutations are missense mutations in the DNA-binding domain, which in addition to loss of canonical p53 activity, frequently confer gain-of-function (GOF) aberrant transcriptional activity through mutant p53 localization to non-canonical genes. GOF phenotypes differ by mutation and cell identity and are reported to include increased proliferation, migration, metabolic reprogramming, and therapy resistance. We found that several recurring p53 mutations exhibit a sex-bias in patients with glioblastoma (GBM). In vitro and in vivo analysis of three mutations, p53R172H, p53Y202C, and p53Y217C revealed sex differences in each mutation′s ability to transform primary mouse astrocytes. p53R172H exhibited a far greater ability to transform female astrocytes than males, p53Y202C transformed both male and female astrocytes with a small male bias, and p53Y217C only exhibited GOF transformation effects in male astrocytes. These phenotypic differences reflect an interaction between sex and GOF mutation to drive unique gene expression patterns in cancer pathways. We found that mutant p53 exhibits sex and mutation specific aberrant genomic localization to the transcriptional start sites of upregulated genes, whose promoter regions were enriched for different sets of transcription factor DNA-binding motifs. Together, our data establish a novel paradigm for sex specific mutant p53 GOF activity in GBM with implications for all cancer.

scholarly journals CBIO-22. p53 GAIN-OF-FUNCTION MUTATIONS DRIVE SEX SPECIFIC EFFECTS ON GLIOMA TUMORIGENESIS

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii20-ii20
Author(s):  
Nathan Rockwell ◽  
Nicole Warrington ◽  
Joshua Rubin
Keyword(s):  
Sex Differences ◽  
Dna Binding ◽  
Target Genes ◽  
Mutant P53 ◽  
Stem Cell Markers ◽  
P53 Mutations ◽  
Gain Of Function ◽  
Male And Female ◽  
Specific Effects

Abstract Sex differences in malignant brain tumors are well-established: Males exhibit greater incidence and poorer survival. Understanding the biology behind these sex differences requires investigation of the pathways known to drive gliomagenesis. The transcription factor TP53 (p53) is one of the most commonly mutated genes in glioblastoma. Most p53 mutations are missense mutations in the DNA-binding domain that lead to the expression of a full length mutant p53 protein. These mutations can endow p53 with oncogenic gains-of-function through aberrant DNA binding and regulation of noncanonical cancer-promoting target genes. Previously, we analyzed patient mutation data and identified six p53 mutations with sex differences in prevalence. In this study, we developed an in vitro mutant p53 glioma model to investigate the sex specific effects of three p53 point mutations: R175H, Y205C, and Y220C (Mm R172H, Y202C, and Y217C respectively). Male and female astrocytes isolated from p53flox/- mouse pups were transduced with a retrovirus expressing mutant p53, followed by a lentivirus expressing CRE recombinase to remove the endogenous WTp53. We then assessed cell proliferation, clonogenicity, and in vivo tumorigenesis in these cells. All three mutations assayed displayed sex differences in proliferation, with male cells overexpressing p53:Y202C and p53:Y217C growing faster than female cells, and female cells overexpressing p53:R172H growing faster than male cells. Male Y202C and Y217C expressing astrocytes also exhibited a trend toward greater clonogenicity compared to female astrocytes. This observation is supported by higher expression of the stem cell markers SOX2 and NESTIN in the male cells. We performed parallel flank injections of male and female astrocytes expressing each mutation or p53 KO. Only male astrocytes expressing p53:Y202C or p53:Y217C and female astrocytes expressing p53:R172H mutation were able to form tumors in vivo. Together, these data support a sex specific gain-of-function phenotype for three different p53 mutations observed in glioma.

scholarly journals CP-31398 Restores DNA-binding Activity to Mutant p53in Vitrobut Does Not Affect p53 Homologs p63 and p73

2004 ◽  
Vol 279 (44) ◽  
pp. 45887-45896 ◽  
Author(s):  
Mark J. Demma ◽  
Serena Wong ◽  
Eugene Maxwell ◽  
Bimalendu Dasmahapatra
Keyword(s):  
Dna Binding ◽  
Mammalian Cells ◽  
P53 Protein ◽  
Binding Activity ◽  
Mutant P53 ◽  
Dependent Manner ◽  
Wild Type ◽  
Dna Binding Activity

The p53 protein plays a major role in the maintenance of genome stability in mammalian cells. Mutations of p53 occur in over 50% of all cancers and are indicative of highly aggressive cancers that are hard to treat. Recently, there has been a high degree of interest in therapeutic approaches to restore growth suppression functions to mutant p53. Several compounds have been reported to restore wild type function to mutant p53. One such compound, CP-31398, has been shown effectivein vivo, but questions have arisen to whether it actually affects p53. Here we show that mutant p53, isolated from cells treated with CP-31398, is capable of binding to p53 response elementsin vitro. We also show the compound restores DNA-binding activity to mutant p53 in cells as determined by a chromatin immunoprecipitation assay. In addition, using purified p53 core domain from two different hotspot mutants (R273H and R249S), we show that CP-31398 can restore DNA-binding activity in a dose-dependent manner. Using a quantitative DNA binding assay, we also show that CP-31398 increases significantly the amount of mutant p53 that binds to cognate DNA (Bmax) and its affinity (Kd) for DNA. The compound, however, does not affect the affinity (Kdvalue) of wild type p53 for DNA and only increasesBmaxslightly. In a similar assay PRIMA1 does not have any effect on p53 core DNA-binding activity. We also show that CP-31398 had no effect on the DNA-binding activity of p53 homologs p63 and p73.

scholarly journals Identification of a druggable protein-protein interaction site between mutant p53 and its stabilizing chaperone DNAJA1

2020 ◽  
pp. jbc.RA120.014749
Author(s):  
Xin Tong ◽  
Dandan Xu ◽  
Rama K. Mishra ◽  
Ryan D Jones ◽  
Leyu Sun ◽  
...  
Keyword(s):  
Small Molecule ◽  
Tp53 Gene ◽  
Dominant Negative ◽  
Site Directed Mutagenesis ◽  
Dominant Negative Effect ◽  
Mutant P53 ◽  
Missense Mutations ◽  
Oncogenic Function

The TP53 gene is the most frequently mutated gene in human cancers, and the majority of TP53 mutations are missense mutations. As a result, these mutant p53 (mutp53) either directly lose wild-type p53 (wtp53) tumor suppressor function or exhibit a dominant negative effect over wtp53. In addition, some mutp53 have acquired new oncogenic function (gain of function). Therefore, targeting mutp53 for its degradation, may serve as a promising strategy for cancer prevention and therapy. Based on our previous finding that farnesylated DNAJA1 is a crucial chaperone in maintaining mutp53 stabilization, and by using an in silico approach, we built 3-D homology models of human DNAJA1 and mutp53R175H proteins, identified the interacting pocket in the DNAJA1-mutp53R175H complex, and found one critical druggable small molecule binding  site in the DNAJA1 glycine/phenylalanine rich region. We confirmed that the interacting pocket in the DNAJA1-mutp53R175H complex was crucial for stabilizing mutp53R175H using a site-directed mutagenesis approach. We further screened a drug-like library to identify a promising small molecule hit (GY1-22) against the interacting pocket in DNAJA1-mutp53R175H complex. The GY1-22 compound displayed an effective activity against DNAJA1-mutp53R175H complex. Treatment with GY1-22 significantly reduced mutp53 protein levels, enhanced Waf1p21 expression, suppressed cyclin D1 expression, and inhibited mutp53-driven pancreatic cancer growth both in vitro and in vivo. Together, our results indicate that the interacting pocket in the DNAJA1-mutp53R175H complex is critical for mutp53’s stability and oncogenic function, and DNAJA1 is a robust therapeutic target for developing the efficient small molecule inhibitors against oncogenic mutp53.

scholarly journals Inhibiting Growth of Human Triple-Negative Breast Cancer Cells by Activating Mutant p53 Protein Alone or in Combination With a Phosphatidylserine-Targeting Antibody

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A1022-A1023
Author(s):  
Yayun Liang ◽  
Rolf A Brekken ◽  
Salman M Hyder
Keyword(s):  
Tumor Suppressor ◽  
Dna Binding ◽  
Blood Vessel ◽  
Cell Viability ◽  
Triple Negative ◽  
Mutant P53 ◽  
Tnbc Cell ◽  
Mcf 7

Abstract Triple-negative human breast cancers (TNBC) lack three proteins commonly targeted by chemotherapy; estrogen and progesterone receptors, and her-2-neu. Most current protocols used to treat TNBC are largely ineffective and aggressive tumors frequently re-emerge, leading to metastasis and patient death. Thus, new therapies for TNBC are needed. Recent studies show that around 80% of TNBC express mutant p53 (mtp53), a functionally defective form of the p53 tumor suppressor protein. If mtp53 is converted into the active wild-type protein (wtp53), tumor suppressor functions are recovered. Most p53 mutations occur in the DNA-binding domain, causing normal regulation of p53 target genes involved in apoptosis, cell-cycle arrest, and angiogenesis to be blocked. This promotes metastasis and renders tumors resistant to chemotherapy. APR-246 is a small-molecule drug that re-activates mtp53 by covalent modification of the DNA-binding core domain of the mutant protein through alkylation of thiol groups and has been shown to reactivate mtp53 and restore p53 function. We examined whether APR-246 could inhibit TNBC growth, both in vitro and in vivo. Cell viability assays and FACS were used to measure in vitro TNBC cell growth and apoptosis respectively, in MDA-MB-231 and MDA-MB-468 cells, with MCF-7 cells (which express wtp53) as controls. Analysis of TNBC growth in vivo was assessed in a mouse model of MDA-MB-231 derived xenografts. Nuclear extracts of APR-246-treated TNBC cells exhibited significantly increased p53 DNA binding compared with untreated cells, indicating that APR-246 converts mtp53 to wtp53 in these cells. APR-246 significantly reduced TNBC cell viability in vitro, but had no effect on normal mammary cells or wtp53-expressing MCF-7 cells. Pro-apoptotic proteins, Bax, p21 and caspase-3 were elevated in APR-246 treated cells, while the cell survival protein Bcl-2 was suppressed. In the xenograft model, animals were given an intravenous (iv) tail vein injection of APR-246 alone (100 mg/kg/day) once tumors reached 100 mm3. A second group received an intraperitoneal (ip) injection of 2aG4 antibody (100 µg/mouse/day), which targets phosphatidylserine and disrupts tumor blood vessel formation. A third group was given both APR-246 and 2aG4 using the same doses above. A control group received antibody C44 (100 µg/mouse/day, ip) and/or PBS (0.1 mL/day, iv). A total of 18 treatments were used. Administration of APR-246 alone or in combination with 2aG4, significantly reduced TNBC tumor growth, as well as two markers of angiogenesis (vascular endothelial growth factor expression and blood-vessel density). APR-246 in combination with 2aG4 completely eradicated almost 20% of the TNBC tumors. We conclude that TNBC is inhibited by APR-246 and 2aG4. Such treatment could represent an effective and innovative means of combating these particularly aggressive and deadly types of cancer.

scholarly journals Effect of 1α,25(OH)2 Vitamin D3 in Mutant P53 Glioblastoma Cells: Involvement of Neutral Sphingomyelinase1

Cancers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3163
Author(s):  
Samuela Cataldi ◽  
Cataldo Arcuri ◽  
Andrea Lazzarini ◽  
Irina Nakashidze ◽  
Francesco Ragonese ◽  
...  
Keyword(s):  
Vitamin D ◽  
Vitamin D3 ◽  
Survival Rates ◽  
P53 Mutation ◽  
Surgical Approaches ◽  
Mutant P53 ◽  
Glioblastoma Cells ◽  
Ceramide Content

Glioblastoma is one the most aggressive primary brain tumors in adults, and, despite the fact that radiation and chemotherapy after surgical approaches have been the treatments increasing the survival rates, the prognosis of patients remains poor. Today, the attention is focused on highlighting complementary treatments that can be helpful in improving the classic therapeutic approaches. It is known that 1α,25(OH)2 vitamin D3, a molecule involved in bone metabolism, has many serendipidy effects in cells. It targets normal and cancer cells via genomic pathway by vitamin D3 receptor or via non-genomic pathways. To interrogate possible functions of 1α,25(OH)2 vitamin D3 in multiforme glioblastoma, we used three cell lines, wild-type p53 GL15 and mutant p53 U251 and LN18 cells. We demonstrated that 1α,25(OH)2 vitamin D3 acts via vitamin D receptor in GL15 cells and via neutral sphingomyelinase1, with an enrichment of ceramide pool, in U251 and LN18 cells. Changes in sphingomyelin/ceramide content were considered to be possibly responsible for the differentiating and antiproliferative effect of 1α,25(OH)2 vitamin D in U251 and LN18 cells, as shown, respectively, in vitro by immunofluorescence and in vivo by experiments of xenotransplantation in eggs. This is the first time 1α,25(OH)2 vitamin D3 is interrogated for the response of multiforme glioblastoma cells in dependence on the p53 mutation, and the results define neutral sphingomyelinase1 as a signaling effector.

scholarly journals Involvement of HECTD1 in LPS-induced astrocyte activation via σ-1R-JNK/p38-FOXJ2 axis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ying Tang ◽  
Mengchun Zhou ◽  
Rongrong Huang ◽  
Ling Shen ◽  
Li Yang ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Astrocyte Activation ◽  
Hect Domain ◽  
P38 Inhibitor ◽  
Ubiquitin Protein Ligase ◽  
Primary Mouse ◽  
Lps Treatment

Abstract Background Astrocytes participate in innate inflammatory responses within the mammalian central nervous system (CNS). HECT domain E3 ubiquitin protein ligase 1 (HECTD1) functions during microglial activation, suggesting a connection with neuroinflammation. However, the potential role of HECTD1 in astrocytes remains largely unknown. Results Here, we demonstrated that HECTD1 was upregulated in primary mouse astrocytes after 100 ng/ml lipopolysaccharide (LPS) treatment. Genetic knockdown of HECTD1 in vitro or astrocyte-specific knockdown of HECTD1 in vivo suppressed LPS-induced astrocyte activation, whereas overexpression of HECTD1 in vitro facilitated LPS-induced astrocyte activation. Mechanistically, we established that LPS activated σ-1R-JNK/p38 pathway, and σ-1R antagonist BD1047, JNK inhibitor SP600125, or p38 inhibitor SB203580 reversed LPS-induced expression of HECTD1, thus restored LPS-induced astrocyte activation. In addition, FOXJ2 functioned as a transcription factor of HECTD1, and pretreatment of primary mouse astrocytes with BD1047, SB203580, and SP600125 significantly inhibited LPS-mediated translocation of FOXJ2 into the nucleus. Conclusions Overall, our present findings suggest that HECTD1 participates in LPS-induced astrocyte activation by activation of σ-1R-JNK/p38-FOXJ2 pathway and provide a potential therapeutic strategy for neuroinflammation induced by LPS or any other neuroinflammatory disorders.

scholarly journals Genome-Wide Binding Analyses of HOXB1 Revealed a Novel DNA Binding Motif Associated with Gene Repression

2021 ◽  
Vol 9 (1) ◽  
pp. 6
Author(s):  
Narendra Pratap Singh ◽  
Bony De Kumar ◽  
Ariel Paulson ◽  
Mark E. Parrish ◽  
Carrie Scott ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Embryonic Stem ◽  
Binding Motif ◽  
Binding Assays ◽  
Histone Marks ◽  
Genome Wide ◽  
Hox Cofactors

Knowledge of the diverse DNA binding specificities of transcription factors is important for understanding their specific regulatory functions in animal development and evolution. We have examined the genome-wide binding properties of the mouse HOXB1 protein in embryonic stem cells differentiated into neural fates. Unexpectedly, only a small number of HOXB1 bound regions (7%) correlate with binding of the known HOX cofactors PBX and MEIS. In contrast, 22% of the HOXB1 binding peaks display co-occupancy with the transcriptional repressor REST. Analyses revealed that co-binding of HOXB1 with PBX correlates with active histone marks and high levels of expression, while co-occupancy with REST correlates with repressive histone marks and repression of the target genes. Analysis of HOXB1 bound regions uncovered enrichment of a novel 15 base pair HOXB1 binding motif HB1RE (HOXB1 response element). In vitro template binding assays showed that HOXB1, PBX1, and MEIS can bind to this motif. In vivo, this motif is sufficient for direct expression of a reporter gene and over-expression of HOXB1 selectively represses this activity. Our analyses suggest that HOXB1 has evolved an association with REST in gene regulation and the novel HB1RE motif contributes to HOXB1 function in part through a repressive role in gene expression.

scholarly journals Gender and Sex Are Key Determinants in Osteoarthritis Not Only Confounding Variables. A Systematic Review of Clinical Data

2021 ◽  
Vol 10 (14) ◽  
pp. 3178
Author(s):  
Matilde Tschon ◽  
Deyanira Contartese ◽  
Stefania Pagani ◽  
Veronica Borsari ◽  
Milena Fini
Keyword(s):  
Health Care ◽  
Sex Differences ◽  
Ex Vivo ◽  
Case Series ◽  
Cartilage Volume ◽  
Review Literature ◽  
Care Needs ◽  
Clinical Pain

Many risk factors for osteoarthritis (OA) have been noted, while gender/sex differences have been understated. The work aimed to systematically review literature investigating as primary aim the relationship between gender/sex related discriminants and OA. The search was performed in PubMed, Science Direct and Web of Knowledge in the last 10 years. Inclusion criteria were limited to clinical studies of patients affected by OA in any joints, analyzing as primary aim gender/sex differences. Exclusion criteria were review articles, in vitro, in vivo and ex vivo studies, case series studies and papers in which gender/sex differences were adjusted as confounding variable. Of the 120 records screened, 42 studies were included. Different clinical outcomes were analyzed: morphometric differences, followed by kinematics, pain, functional outcomes after arthroplasty and health care needs of patients. Women appear to use more health care, have higher OA prevalence, clinical pain and inflammation, decreased cartilage volume, physical difficulty, and smaller joint parameters and dimensions, as compared to men. No in-depth studies or mechanistic studies analyzing biomarker differential expressions, molecular pathways and omic profiles were found that might drive preclinical and clinical research towards sex-/gender-oriented protocols.

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