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p53 mutation heterogeneity in cancer
Abstract
The p53 gene is inactivated in about 50% of human cancers and the p53 protein is an essential component of the cell response induced by genotoxic stresses such as those generated by radiotherapy or chemotherapy. It is therefore highly likely that these alterations are an important component in tumor resistance to therapy. The particular characteristics of these alterations, 80% of which are missense mutations leading to functionally heterogeneous proteins, make p53 a unique gene in the class of tumor suppressor genes. A considerable number of mutant p53 proteins probably have an oncogenic activity per se and therefore actively participate in cell transformation. The fact that the apoptotic and antiproliferative functions of p53 can be dissociated in certain mutants also suggests another level of complexity in the relationships between p53 inactivation and neoplasia.
... This is the target for 90% ofp53 mutations found in human cancers [111]. The tetramerisation domain consists of a beta-strand, followed by an alpha-helix necessary for dimerisation [112]. ...
... This selection to maintain mutant p53 in tumour cells is believed to be required for both a dominant negative activity, to inhibit wild-type p53 expressed by the remaining allele, and for gain of function that transforms mutant p53 into a dominant oncogene [116][117][118]. Although the majority of mutations are localised to the DNA binding domain ofp53, mutant p53 proteins exhibit structural, biochemical and biological heterogeneity [111]. Certain p53 mutants are able to transactivate genes involved in proliferation or cell survival and these sites differ from those bound by wild-type p53 [111]. ...
... Although the majority of mutations are localised to the DNA binding domain ofp53, mutant p53 proteins exhibit structural, biochemical and biological heterogeneity [111]. Certain p53 mutants are able to transactivate genes involved in proliferation or cell survival and these sites differ from those bound by wild-type p53 [111]. Other p53 mutants are able to interact directly with cellular proteins involved in tumour progression [119]. ...
p>Inactivating mutations in the TP53 gene are an infrequent event in tumours derived from cells of neural crest origin. The survival of cells from these tumours is often dependent upon their expression of PAX3, an embryologically expressed transcription factor that is essential for the suppression of spontaneous p53-dependent apoptosis in the developing neural crest. I hypothesized that PAX3 causes repression of p53 function in tumours of neural crest origin.
I show that PAX3 is required to prevent spontaneous p53-dependent apoptosis in melanoma cells using siRNA. Knockdown of PAX3 expression causes a loss of cell viability of over 90% and induction of the pro-apoptotic protein BAX. This is in contrast to induction of the cyclin dependent kinase inhibitor p21<sup>WAF-1</sup> and cell cycle arrest in response to HDM2 knockdown. Furthermore, I confirm the importance of PAX3 in the survival of neuroblastoma.
In a model system PAX3 suppresses p53-dependent transcription from promoters including BAX and HDM2 but, importantly, not WAF-1 . Mutagenesis studies show that this suppression is dependent upon the integrity of PAX3 as an activating transcription factor. PAX3 expression causes a reduction in p53 protein levels by up to 90% and an increase in p53 turn-over, which is not due to classical proteasomal degradation.
My data clearly demonstrates a critical role for the PAX3 oncoprotein in the inhibition of p53-dependent pro-apoptotic pathways in neural crest derived tumours. Furthermore, I have developed a cell based screening assay to identify compounds that inhibit PAX3 function in melanoma cells. Such compounds will be used as tools to study the mechanism of p53 functional inhibition by PAX3 in future work and may also have therapeutic potential.</p
... It affects many cell functions involved in the stress response-modifies cellular metabolism, influences autophagy and apoptotic processes, and acts as a multipotent transcription factor inducing or inhibiting the transcription of numerous target genes of this protein [18,19]. An example of two areas (cytoplasmic and nuclear) of p53 activity is the effect on the induction of apoptosis in cell cultures exposed to genotoxic stress [20]. p53 forms a complex with the anti-apoptotic Bcl-xL protein, induces oligomerization and activation of the pro-apoptotic Bak protein, and increases the release of cytochrome c from the mitochondria. ...
... p53 forms a complex with the anti-apoptotic Bcl-xL protein, induces oligomerization and activation of the pro-apoptotic Bak protein, and increases the release of cytochrome c from the mitochondria. The p53 protein significantly increases apoptosis as early as 30 min after exposure of cells to a genotoxic agent [20,21]. The effect in the cell nucleus is observed after 2 h from exposure to stress and is manifested by intensifying the transcription of factors enhancing apoptosis of the mitochondrial pathway [22]. ...
... Mutations of this gene are found on average in more than 55% of cases of human cancers, and even more in some types of cancer, e.g. in pancreatic cancer and advanced ovarian cancer-75% and 96% (respectively) of clinical cases show the TP53 mutation [26]. Over 80% of the described mutations of this gene are missense mutations that lead to increased expression of the p53 protein differing from the wild-type protein with one amino acid [20]. The protein product of such a mutant TP53 gene loses its physiological function of inhibiting the cell division cycle and inducing apoptosis (functions of the tumor suppressor protein). ...
Background
The p53 protein is a transcription factor for many genes, including genes involved in inhibiting cell proliferation and inducing apoptosis in genotoxically damaged and tumor-transformed cells. In more than 55% of cases of human cancers, loss of the essential function of p53 protein is found. In numerous reports, it has been shown that small molecules (chemical compounds) can restore the suppressor function of the mutant p53 protein in tumor cells. The aim of this study was to evaluate the potential anticancer activity of three newly synthesized olivacine derivatives.
Methods
The study was performed using two cell lines—CCRF/CEM (containing the mutant p53 protein) and A549 (containing a non-mutant, wild-type p53 protein). The cells were incubated with olivacine derivatives for 18 h and then assays were carried out: measurement of the amount of p53 and p21 proteins, detection of apoptosis, cell cycle analysis, and rhodamine 123 accumulation assay (evaluation of P-glycoprotein inhibition). Multiple-criteria decision analysis was used to compare the anticancer activity of the tested compounds.
Results
Each tested compound caused the reconstitution of suppressor activity of the p53 protein in cells with the mutant protein. In addition, one of the compounds showed significant antitumor activity in both wild-type and mutant cells. For all compounds, a stronger effect on the level of the p53 protein was observed than for the reference compound—ellipticine.
Conclusions
The observed effects of the tested new olivacine derivatives (pyridocarbazoles) suggest that they are good candidates for new anticancer drugs.
... In recent years, an increasing number of studies have recognized the importance of TP53 in tumors (25)(26)(27). p53 is a transcription factor that stabilizes genotoxic stress and induces the transcription of genes involved in cell cycle arrest, apoptosis and metabolism, thereby acting as a tumor suppressor (22). In general, tumor suppressors have a loss-of-function or deletion mutation in cancers (28). ...
... The frequency of complex karyotypes in TP53-positive patients is significantly higher than that in TP53-negative patients (33). At the molecular level, since TP53 mutations mostly occur in exons of the DNA binding domain, the arginine residue of the p53 protein is considered to be a mutation hot spot, which can affect DNA binding and change the activity of the mutant protein (25,26,34). TP53 is associated with chemosensitivity, mainly by affecting the patient's chemical tolerance or overall survival rate (27). ...
Myeloid sarcoma (MS) is a rare extramedullary tumor mass that carries a high risk of progression to acute myeloid leukemia (AML), and patients with MS are commonly treated with the AML regimen. However, MS is frequently misdiagnosed due to its lack of clinical specificity. Patients with MS who harbor tumor protein p53 (TP53) mutations and complex karyotypes are considered to have a poorer prognosis. The present study reports a case of lymph node MS with TP53 (V173G)-related myelodysplastic syndrome (MDS). The mass was first considered to be a lymphoma and treated as such. However, following immunohistochemical analysis, which revealed cells positive for CD43, myeloperoxidase and CD117, the patient was later diagnosed with MS combined with MDS. The patient went into complete remission after the first cycle of chemotherapy, and showed a decrease in platelet, red blood cell and white blood cell counts following the second cycle of chemotherapy. After the third chemotherapy, agranulocytosis occurred, leading to refractory pneumonia and eventually death due to respiratory failure. MS with TP53-related MDS has a low incidence rate, a poor prognosis and a short survival time. The clinical manifestations of MS are non-specific and easy to misdiagnose, leading to delayed diagnosis and treatment, and ultimately worsening the prognosis of the patients. Therefore, a lymph node biopsy should be performed as soon as possible for patients with lymph node enlargement, and early treatment should be carried out to prolong the survival period.
... The Zn particles are cofactors of the superoxide dismutase compounds, which forestall the beginning and movement of tumors through cell security against substances that cause the development of free radical oxygen species. A low amount of Zn was in mentioned in literature for tumors of liver, kidney, uterus, lung prostate, stomach, testis, thyroid and in esophageal squamous cell cancer [35][36][37]. These facts suggest that reduced level of Zn content in tumors is presumably one of the elements in the etiology of harmful change of various tissues, because Zn insufficiency has been connected to severe shortage of immune and interruption in T-Cell function. ...
... An increasing amount of Cd in humans may interference with the different metabolic methods of Zn and frequently treated to decrease the adverse effect of Cd [39]. Protein bound sulfhydryl groups and Cd depletes glutathione subsequent improved the production of superoxide ion hydrogen peroxide and hydroxyl radicals may inhibit with key strides in the restoring of damaged DNA modulations [37]. Cancer patients have insufficient protein consumptions and energy so starvation is chief problem for cancerous patients. ...
Background
Ewing’s sarcoma is a very rare type of cancerous tumor that grows in bones or the soft tissue around the bones, such as cartilage or the nerves. It frequently affects the people at the age of 10 to 20 years and has elevated rate of being cured.
Methods
Assessment of essential trace [zinc (Zn)] and toxic [cadmium (Cd)] elements in biological samples (scalp hair and blood) of Ewing sarcoma patients (n=87 age ranging 07-19 years) residents of various cities of Pakistan was done. For comparative study, the biological samples of age matched healthy (referent) subjects (n= 62) were also analyzed for selected metals. The matrices of biological samples was oxidized with the help of HNO3 (65%) and H2O2 (30%) mixture at the ratio of 2:1 with the aid of microwave oven. The oxidized biological samples were subjected to atomic absorption spectrometry for their determination.
Result
The Zn contents in the scalp hair and blood samples of different types of Ewing sarcoma patients were found to be lower, in the range of (45.9- 141.2 µg/g) and (0.65- 3.12 mg/l), respectively than the biological samples of referent subjects (246- 265 µg/g) and (6.40- 7.25 mg/l), respectively. Whilst, the Cd concentrations in the scalp hair and blood samples of different types of Ewing sarcoma patients were found to be higher, in the range of (2.70- 5.60 µg/g) and (2.46- 5.64 µg/l), respectively than the biological samples of controls (1.49- 1.79 µg/g) and (1.52- 1.90 µg/l), respectively. The biochemical parameters including hemoglobin % and RBC counts were significantly lower in patients than referents (p<0.001), whereas WBC counts was alarmingly higher in patients than referents (p<0.001).
Conclusion
The resulted data will be helpful to treat patients of Ewing sarcoma with mineral supplement contains essential trace element (Zn) in recommended dose to further increase their survival rate.
... The role of elements is important in human life for normal functions of the human body system. The changes in the levels of essential and toxic elements in the human body tissues produce adverse impacts on the physiological and pathological functions; however, the abundance of one element interferes with the task of another to pharmacologically active dose and life threating [7,29]. ...
... These facts suggest that reduced levels of Zn in tumors are presumably one of the elements in the etiology of harmful change of various tissues, because Zn insufficiency has been connected to severe shortage of immune and interruption in T cell function. P53 inactivation has been caused by deficiency of Zn, a tumor suppressor protein, which has been related with different diseases [29]. In Table 6, the Zn/Cd mole ratio in control and different types of female breast cancer was presented. ...
Breast cancer is the most familiar solid tumor analyzed in women. Trace elements have critical roles in cancer biology. In this research, the relationship between carcinogenic element, cadmium (Cd), and anti-carcinogenic elements, zinc (Zn), in the scalp hair and blood samples of four stages of female breast cancer patients was studied. We have determined the essential trace (Zn) and toxic (Cd) elements, in biological samples (scalp hair and blood) of female breast cancer (n = 96 age ranging 22–35 years), residents of various cities of Pakistan. For comparative study, the biological samples of age-matched healthy (referent) subjects (n = 115) were also analyzed for selected metals. The validity and accuracy of the methodology were checked by using certified reference materials of biological referent materials (human hair (BCR 397) and ClinCheck lyophilized blood). The mean concentrations of Cd were found to be 3- to fourfold significantly higher in the scalp hair and blood samples of female breast patients as compared to referents, while reverse results were obtained in the case of Zn (p > 0.001). The observed data shows the significant effect of carcinogenic (Cd) and their balance towards the anti-carcinogenic (Zn) in humans.
... TP53 is the most mutated gene in human cancer [90], and, at some point during tumor progression, about 50% of cancers acquire a p53 mutation [91]. Modifications have been found in every region of the protein [90], but only some of the most frequently occurring mutations have seen their contribution to cancer progression thoroughly studied. ...
... Modifications have been found in every region of the protein [90], but only some of the most frequently occurring mutations have seen their contribution to cancer progression thoroughly studied. While most tumor suppressor genes are inactivated by mutations, leading to loss of protein synthesis, missense mutations represent 80% of the alterations for TP53 [91]. Frameshift and nonsense mutations, which normally result in the loss of p53 expression, are possible in any case. ...
The cystine/glutamate antiporter xCT is a tumor-associated antigen that has been newly identified in many cancer types. By participating in glutathione biosynthesis, xCT protects cancer cells from oxidative stress conditions and ferroptosis, and contributes to metabolic reprogramming, thus promoting tumor progression and chemoresistance. Moreover, xCT is overexpressed in cancer stem cells. These features render xCT a promising target for cancer therapy, as has been widely reported in the literature and in our work on its immunotargeting. Interestingly, studies on the TP53 gene have revealed that both wild-type and mutant p53 induce the post-transcriptional down modulation of xCT, contributing to ferroptosis. Moreover, APR-246, a small molecule drug that can restore wild-type p53 function in cancer cells, has been described as an indirect modulator of xCT expression in tumors with mutant p53 accumulation, and is thus a promising drug to use in combination with xCT inhibition. This review summarizes the current knowledge of xCT and its regulation by p53, with a focus on the crosstalk of these two molecules in ferroptosis, and also considers some possible combinatorial strategies that can make use of APR-246 treatment in combination with anti-xCT immunotargeting.
... These combinations may allow MEG3 to perform different functions and need to be further con rmed. It has been reported that the expression of P53 in tumors is usually associated with missense mutations, ie, a gene encoding P53 is mutated to synthesize a mutant of P53 (72). Most mutant P53 have proto-oncogene activity and can promote malignant transformation of cells. ...
... Most mutant P53 have proto-oncogene activity and can promote malignant transformation of cells. However, some mutant P53 have a pro-apoptotic ability to exert a tumor suppressor function (72). Therefore, based on our results, it can be speculated that human liver cancer stem cells isolated from Huh7 cells may express wild-type P53 or some mutant P53, and this mutant may contain R175H, P275S, L344P. ...
Background: MEG3 is abnormally down-regulated in most tumors and inhibits tumorigenesis.
Methods: Gene infection, Western blotting and tumorigenesis test in vitro and in vivo were performed to analyze the signaling pathway.
Results: MEG3 increased the loading of P300 and RNA polymerase II onto the promoter regions of P53. Notably, MEG3 increased the methylation of histone H3 at lysine 27 through increasing the interplay between PRC2 and histone H3. Furthermore, MEG3 inhibited the expression of TERT by increasing the H3K27me3 and decreasing the loading of RNA pol Ⅱ in TERT promoter regions. Moreover, MEG3 inhibit the activity of telomerase by reducing the binding of TERT to TERC competitively. In addition, MEG3 also increased the TERRA through reducing DNA methyltransferase DNMT3b binding to the promoter regions of TERRA competitively. Therefore, the interaction between TERC and TERT was competitively attenuated by increasing the interaction between TERRA and TERT, which inhibited the activity of telomerase in hLCSCs.In particular, MEG3 shortened the length of telomere by blocking the formation of complex maintaining telomere length(POT1-Exo1-TRF2-SNM1B) and decreasing the binding of the complex to telomere competitively, which was caused by increasing the interplay between P53 and HULC in hLCSCs.Strikingly, MEG3 inhibited the growth in vitro and in vivo of hLCSCs by reducing the activity of telomerase and attenuating telomeric repeat binding factor 2(TRF2).
Conclusions: our results demonstrated MEG3 inhibits the occurrence of human liver cancer and these findings provide an important insight into the prevention and treatment of human liver cancer.
... These combinations may allow MEG3 to perform different functions and need to be further con rmed. It has been reported that the expression of P53 in tumors is usually associated with missense mutations, ie, a gene encoding P53 is mutated to synthesize a mutant of P53 (72). Most mutant P53 have proto-oncogene activity and can promote malignant transformation of cells. ...
... Most mutant P53 have proto-oncogene activity and can promote malignant transformation of cells. However, some mutant P53 have a pro-apoptotic ability to exert a tumor suppressor function (72). Therefore, based on our results, it can be speculated that human liver cancer stem cells isolated from Huh7 cells may express wild-type P53 or some mutant P53, and this mutant may contain R175H, P275S, L344P. ...
Background: MEG3 is abnormally down-regulated in most tumors and inhibits tumorigenesis.
Methods: Gene infection, Western blotting and tumorigenesis test in vitro and in vivo were performed to analyze the signaling pathway.
Results: MEG3 increased the loading of P300 and RNA polymerase II onto the promoter regions of P53. Notably, MEG3 increased the methylation of histone H3 at lysine 27 through increasing the interplay between PRC2 and histone H3. Furthermore, MEG3 inhibited the expression of TERT by increasing the H3K27me3 and decreasing the loading of RNA pol Ⅱ in TERT promoter regions. Moreover, MEG3 inhibit the activity of telomerase by reducing the binding of TERT to TERC competitively. In addition, MEG3 also increased the TERRA through reducing DNA methyltransferase DNMT3b binding to the promoter regions of TERRA competitively. Therefore, the interaction between TERC and TERT was competitively attenuated by increasing the interaction between TERRA and TERT, which inhibited the activity of telomerase in hLCSCs.In particular, MEG3 shortened the length of telomere by blocking the formation of complex maintaining telomere length(POT1-Exo1-TRF2-SNM1B) and decreasing the binding of the complex to telomere competitively, which was caused by increasing the interplay between P53 and HULC in hLCSCs.Strikingly, MEG3 inhibited the growth in vitro and in vivo of hLCSCs by reducing the activity of telomerase and attenuating telomeric repeat binding factor 2(TRF2).
Conclusions: our results demonstrated MEG3 inhibits the occurrence of human liver cancer and these findings provide an important insight into the prevention and treatment of human liver cancer.
... XYL from sweeteners, MPG from flavor enhancers, and SS among preservatives showed the highest affinity. P53 protein is crucial for interrupting the cell cycle and obstructing cancer development, so inactivation of the protein is associated with many malignancies (Joerger and Fersht, 2016;Soussi and Lozano, 2005;Ueda et al., 1995). In a study on the inhibition of p53 protein, binding constants were determined between − 5.01 to − 7.14 kcal/mol, and a stable interaction was emphasized (Kalungi et al., 2023). ...
... Several polymorphisms of TP53 have been studied. The SNP p.R72P (rs1042522) of the TP53 gene is one of the most studied polymorphisms of TP53 gene [8,13]. It is located in exon and characterized by the presence of an arginine (R) or a proline (P) in position 72 [7]. ...
Introduction: The TP53 and CHEK2 genes have been described as breast cancer susceptibility genes and some of their polymorphisms have been associated with an increased risk of breast cancer in certain populations.Aim: The objective of this study was to investigate the p.R72P and PIN3 Ins16bp (TP53) polymorphisms and the I157T (CHEK2) mutation developping of breast cancer. Methods: This case-control study had enrolled 144 participants including 65 cases (breast cancer patients) and 79 controls (women without breast abnormalities) in the city of Ouagadougou in Burkina Faso. The DNA was extracted using the method of “salting out” and the genotyping of polymorphisms was performed by ASO-PCR (Allele Specific Oligonucleotides - Polymerase Chain Reaction), conventional PCR and PCR-RFLP (Polymerase Chain Reaction - Restriction Fragment Length Polymorphism) techniques. Results: The heterozygous genotype (RP) of the p.R72P polymorphism of TP53 gene was in the majority in cases (73.85%) and controls (73.42%). Regarding to the PIN3 Ins16bp polymorphism of TP53 gene, the homozygous wild type (A1A1) was the most represented in both cases (53.85%) and controls (60.76%). Concerning the I157T mutation of CHEK2 gene, only one (01) patient was homozygous mutant (TT) and no controls had the mutation. This study found no association between these polymorphisms and the risk of breast cancer occurrence (p.R72P (OR=0.96; 95%IC (0.59-1.56); p=0.471), PIN3 Ins16bp (OR= 1.1; 95%IC (0.61-1.98); p=0.420)). Conclusion: This study showed that the P allele of the p.R72P polymorphism and the wild-type allele (A1) of the PIN3 Ins16bp polymorphism were in the majority. The I157T mutation was very rare. These polymorphisms were not associated with the risk of developing breast cancer in this study.
... Several polymorphisms of TP53 have been studied. The SNP p.R72P (rs1042522) of the TP53 gene is one of the most studied polymorphisms of TP53 gene [8,13]. It is located in exon and characterized by the presence of an arginine (R) or a proline (P) in position 72 [7]. ...
Introduction: The TP53 and CHEK2 genes have been described as breast cancer susceptibility genes and some of their polymorphisms have been associated with an increased risk of breast cancer in certain populations.
Aim: The objective of this study was to investigate the p.R72P and PIN3 Ins16bp (TP53) polymorphisms and the I157T (CHEK2) mutation developping of breast cancer.
Methods: This case-control study had enrolled 144 participants including 65 cases (breast cancer patients) and 79 controls (women without breast abnormalities) in the city of Ouagadougou in Burkina Faso. The DNA was extracted using the method of “salting out” and the genotyping of polymorphisms was performed by ASO-PCR (Allele Specific Oligonucleotides - Polymerase Chain Reaction), conventional PCR and PCR-RFLP (Polymerase Chain Reaction - Restriction Fragment Length Polymorphism) techniques.
Results: The heterozygous genotype (RP) of the p.R72P polymorphism of TP53 gene was in the majority in cases (73.85%) and controls (73.42%). Regarding to the PIN3 Ins16bp polymorphism of TP53 gene, the homozygous wild type (A1A1) was the most represented in both cases (53.85%) and controls (60.76%). Concerning the I157T mutation of CHEK2 gene, only one (01) patient was homozygous mutant (TT) and no controls had the mutation. This study found no association between these polymorphisms and the risk of breast cancer occurrence (p.R72P (OR=0.96; 95%IC (0.59-1.56); p=0.471), PIN3 Ins16bp (OR= 1.1; 95%IC (0.61-1.98); p=0.420)).
Conclusion: This study showed that the P allele of the p.R72P polymorphism and the wild-type allele (A1) of the PIN3 Ins16bp polymorphism were in the majority. The I157T mutation was very rare. These polymorphisms were not associated with the risk of developing breast cancer in this study.
... In this regard, the altered activities of the GOF-mutants are often dictated by the proteins available to interact with it in the intracellular milieu, which in turn can be both context and tumor cell type dependent. In addition, different GOF point mutations can result in altered conformation of mutant-P53 potentially changing the extent and array of GOFs [12]. Therefore, it is critical to analyse the most prevalent GOF-P53 point mutations in individual cancer types, identify their probable interacting partners and elucidate their potential biological function. ...
Background
Non-small cell lung carcinoma (NSCLC) is the most common cause of cancer-associated deaths worldwide. Though recent development in targeted therapy has improved NSCLC prognosis, yet there is an unmet need to identify novel causative factors and appropriate therapeutic regimen against NSCLCs.
Methods and results
In this study, we identify key molecular factors de-regulated in NSCLCs. Analyze their expression by real-time PCR and immunoblot; map their localization by immuno-fluorescence microscopy. We further propose an FDA approved drug, chloroquine (CQ) that affects the function of the molecular factors and hence can be repurposed as a therapeutic strategy against NSCLCs. Available NSCLC mutation data reflects a high probabilistic chance of patients harboring a p53 mutation, especially a gain of function (GOF)-R273H mutation. The GOF-P53 mutation enables the P53 protein to potentially interact with non-canonical protein partners facilitating oncogenesis. In this context, analysis of existing transcriptomic data from R273H-P53 expressing cells shows a concomitant up-regulation of Yes-associated protein (YAP) transcriptional targets and its protein partner TEAD1 in NSCLCs, suggesting a possible link between R273H-P53 and YAP. We therefore explored the inter-dependence of R273H-P53 and YAP in NSCLC cells. They were found to co-operatively regulate NSCLC proliferation. Genetic or pharmacological inhibition of YAP and GOF-P53 resulted in sensitization of NSCLC cells. Further analysis of pathways controlled by GOF-P53 and YAP showed that they positively regulate the cellular homeostatic process- autophagy to mediate survival. We hence postulated that a modulation of autophagy might be a potent strategy to curb proliferation. In accordance to above, autophagy inhibition, especially with the FDA-approved drug- chloroquine (CQ) resulted in cytoplasmic accumulation and reduced transcriptional activity of GOF-P53 and YAP, leading to growth arrest of NSCLC cells.
Conclusion
Our study highlights the importance of GOF-P53 and YAP in NSCLC proliferation and proposes autophagy inhibition as an efficient strategy to attenuate NSCLC tumorigenesis.
... In our study, the positivity of TP53 expression was only 25.2%. The result obtained was below the corresponding values found in the literature, which indicate values between 50% and 70% derived from neoplasms and different techniques for evaluating expression (6,(28)(29)(30)(31) . The discrepant finding may be related to the increasing technological sophistication of diagnostic methods with greater sensitivity, such as those based on the Polymerase Chain Reaction (PCR) (20,21) . ...
Introduction: In Brazil, 8,000 new cases of childhood cancer are estimated each year, whose causes are still little known, although some
have genetically determined factors. Approximately 70% of human cancers have alterations in the TP53 gene, which encodes the protein
responsible for inhibiting the disordered growth of cells exposed to injuries. However, the frequency of alterations in the expression of TP53
in childhood cancers in Brazil remains poorly known.
Objective: To evaluate the expression of TP53 gene in patients with childhood cancer in northeastern of Santa Catarina, Brazil. Materials
and Methods: Retrospectively, 282 patients diagnosed with cancer between 2005 and 2015 in Joinville were included. TP53 expression was
evaluated by immunohistochemistry using a score based on the intensity and percentage of stained cells.
Results: The p53 protein was positive in 25.2% of cases, with no difference between sexes. Considering the five main groups of tumors in
the sample, the expression was positive in 31.8%, 27.3%, 20%, 17.2% and 5.9% of lymphomas, nephroblastomas, neuroblastomas, tumors
of the Central Nervous System and leukemias, respectively.
Conclusion: The prevalence of TP53 expression was evaluated in different childhood cancers in the northeastern of Santa Catarina.
Positivity was higher among lymphomas and lower in leukemias, but with no significant difference among the five most frequent tumors.
Further studies that allow correlation with aggressiveness and disease evolution are required.
... More than 50 % of human cancers are shown to contain mutations in TP53 gene and most commonly the mutations are missense mutations which is a single nucleotide substituted by another. [5] The detection of TP53 mutation by liquid biopsy techniques presents the potential in replacing invasive, painful and costly tissue biopsies in cancer diagnosis and changing the way of monitoring human malignancies at high resolution. [6] Thus, using mutant TP53 gene as a potential liquid biopsy-based cancer biomarker presents several advantages, including non-invasive, non-localized and remotely sampling from the site of pathology. ...
Mutant TP53 has been demonstrated to be a promising biomarker as it is the most frequently mutated gene in human cancer. However, quantifying TP53 mutation presents two unique challenges: a broad concentration range and ultralow abundance. This study reports on the development of an ultrasensitive and highly‐selective electrochemical biosensor for the rapid detection of TP53 mutation in blood. TP53 mutation analysis is based on a network of gold‐coated magnetic nanoparticles modified with probe DNA (DNA−Au@MNPs). The DNA−Au@MNPs biosensor can rapidly and selectively detect TP53 gene with a single‐base mutation at a high background of wild‐type TP53 gene in blood. Moreover, the sensor presents a broad dynamic range of 1 aM to 10 nM for the mutant TP53 gene with the lowest detected concentration of 1 aM and significantly reduced response time of 20 min. Therefore, this biosensor demonstrates the potential to be used for minimally invasive analysis of cancer.
... Indeed, the p53 protein, considered as a "guardian of the genome", plays many major roles in cell function [14]. The SNP p.R72P (rs1042522) of the TP53 gene is one of the most studied polymorphisms of TP53 [9,15]. It is located in exon 4 characterized by the presence of an arginine (R) or a proline (P) in position 72 [16]. ...
Background: The TP53 and CHEK2 genes have been described as breast cancer susceptibility genes and some of their polymorphisms have been associated with an increased risk of breast cancer in certain populations. The objective of this study was to investigate the p.R72P and PIN3 Ins16bp (TP53) polymorphisms and the I157T (CHEK2) mutation.
Methods and results: This case-control study had enrolled 144 participants including 65 cases (breast cancer patients) and 79 controls (women without breast abnormalities). The genotyping of polymorphisms was performed by ASO-PCR (Allele Specific Oligonucleotides - Polymerase Chain Reaction) and PCR-RFLP (Polymerase Chain Reaction - Restriction Fragment Length Polymorphism) techniques.
The heterozygous mutated genotype (RP) of the p.R72P polymorphism was in the majority in cases (73.85%) and controls (73.42%). As for the PIN3 Ins16bp polymorphism, the homozygous wild type (A1A1) was the most represented in both cases (53.85%) and controls (60.76%). Concerning the I157T mutation, only one (01) patient was homozygous mutant (TT) and no controls had the mutation. This study found no association between these polymorphisms and the risk of breast cancer occurrence (p.R72P (OR=0.96; 95%IC (0.59-1.56); p=0.471), PIN3 Ins16bp (OR= 1.1; 95%IC (0.61-1.98); p=0.420)).
Conclusion: This study showed that the mutated allele of the p.R72P polymorphism versus the wild-type allele of the PIN3 Ins16bp polymorphism were in the majority. The I157T mutation was very rare. These polymorphisms were not associated with the risk of developing breast cancer.
... PURPL, a lncRNA downregulated in RBPMS knockout clones, regulates p53 unction in many cancer types [77]. As p53 gain-of-function (GOF) mutants have been reported in many cancer types [78][79][80], it could be interesting to investigate if increasing PURPL levels could reduce the p53 levels in cisplatin-resistant ovarian cancer cells. The role of the other three deregulated ncRNAs upon RBPMS knockout-LINC01036 (upregulated), LOC101927789 (downregulated), and LOC105377329 (downregulated)-have not yet been studied in cancer. ...
Citation: Rabelo-Fernández, R.J.; Santiago-Sánchez, G.S.; Sharma, R.K.; Roche-Lima, A.; Carrion, K.C.; Rivera, R.A.N.; Quiñones-Díaz, B.I.; Rajasekaran, S.; Siddiqui, J.; Miles, W.; et al. Reduced RBPMS Levels Promote Cell Proliferation and Decrease Cisplatin Sensitivity in Ovarian Cancer Cells. Int. J. Mol. Sci. 2022, 23, 535. https://doi.
... TP53, one of the most well-known tumor-suppressive genes, controls cell proliferation via the regulation of the cell cycle and apoptosis (21,22). A mutation in TP53 is reported to reduce the anti-proliferative and apoptotic functions of p53 and occasionally even exert an oncogenic function (37). In the present study, AGR2 downregulation increased the phosphorylation of p53 Ser15 and decreased malignant potential in the TP53-wild-type cell lines TE15, TE2 and U2OS, but not in the TP53-mutant cell line TE5 because the phosphorylation of p53 is frequently induced without AGR2 expression in p53-mutant cells. ...
Anterior gradient 2 (AGR2) reportedly promotes tumor growth and has an unfavorable impact on survival in several cancers. However, no comprehensive functional analysis of AGR2 in esophageal squamous cell carcinoma (ESCC) has been performed. In the present study, the function and clinical significance of AGR2 were examined using ESCC cell lines and clinical samples. AGR2 was upregulated in EC tissue and ESCC cell lines. The downregulation of AGR2 suppressed cell proliferation and increased the proportion of G2/M‑phase cells and phosphorylation of p53 in TP53‑wild‑type ESCC and osteosarcoma cells. However, these changes were not observed in TP53‑mutant ESCC cells. In addition, immunohistochemistry results demonstrated that high AGR2 and low p53 expression levels in ESCC tissues were correlated with a worse prognosis. These results suggested that although AGR2 enhanced cell proliferation by inhibiting p53 phosphorylation in TP53‑wild‑type ESCC, the same mechanism did not regulate cell functions in TP53‑mutant ESCC. Thus, AGR2 served an important role in ESCC progression and might be a useful prognostic marker in patients with TP53‑wild‑type ESCC.
... The most well studied mutations in TP53 occur in the DNA binding domain of the protein. Mutations in this region impair the ability of p53 to bind to its target genes and result in the accumulation of mutant p53 (Soussi and Lozano, 2005). Mouse models expressing mutant p53 develop more aggressive cancers and increased metastasis compared to mice lacking p53 all together (Lang et al., 2004;Olive et al., 2004). ...
The tumor suppressor gene TP53 is the most frequently mutated gene in cancer and plays a key role in mediating several processes that are critical for preventing tumor formation and progression. Known as the guardian of the genome, p53 regulates hundreds of genes involved in various pathways such as apoptosis, cell cycle arrest and senescence. In recent years, the role of p53 in metabolism, redox state and ferroptosis has begun to emerge. Our lab has identified an African-specific polymorphic variant of p53 that encodes a serine residue instead of a proline at amino acid 47 (hereafter S47) and predisposes carriers to cancer. The S47 variant is impaired for tumor suppression and ferroptosis, and S47 cells have an altered redox state. We sought to use the tumor prone S47 model as a tool to better understand the role of p53 in tumor suppression. Our results demonstrate that mice carrying the S47 variant have greater metabolic efficiency compared to those with WT p53, along with increased mTOR activity. This difference in mTOR stems from an impaired protein-protein interaction that occurs in S47, ultimately due to a difference in cellular redox state. We next identified PLTP as a p53 target gene that shows decreased transactivation in the S47 variant and mediates ferroptosis resistance by enhancing lipid storage in HepG2 cells. Taken together, this work sheds light on the emerging roles p53 plays in tumor suppression, metabolism and ferroptosis. It also provides a better understanding of an ethnic genetic variant of p53. We expect this work will enable better personalized medicine approaches and therapeutic options for people who carry this variant.
... Mutations in the guardian of the genome, p53, occur in almost 50% of all cancers 7 . In CLL, mutations of this gene have been found in about 4-37% of patients [8][9][10] , some of which are associated with poor prognosis 11,12 . ...
Background: Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults. The MDM2 and p53 are interacting proteins that play crucial roles in cell biology. Genetic variations of p53 and MDM2 (p53 codon 72 and MDM2 SNP30) have been identified in many cancers including CLL.
Materials and Methods: In this study, we sought to find the impact of two SNPs of p53 and MDM2 in the pathogenesis of CLL. A total of 100 CLL patients and 102 healthy controls were recruited. Genomic DNA was extracted, and genotyping was performed using the PCR-RFLP method. The allele and genotype associations were analyzed using the χ2 test. The gene-gene interaction analysis was studied using GMDR v0.9.
Results: Our study found the absence of a significant difference between CLL patients and controls related to the allelic frequencies or genotypic distributions for both MDM2 SNP309 and p53 codon72. A significantly higher frequency of p53 C allele was found in patients with a disease duration of more than 36 compared to those less than 36 months. However, GMDR analysis suggests genetic interaction between the genes under study.
Conclusion: Our findings indicated each polymorphism of p53 codon72 and MDM2 (SNP309) was not a risk factor for CLL but the p53 C allele could be associated with the disease duration. Besides, the interaction between p53/MDM2 genotypes may confer susceptibility to CLL. Our study could be useful in genetic association studies of CLL and the role of gene-gene interactions in the susceptibility to the disease.
... De façon intéressante, plus de 25% des mutations de p53 dans les cancers humains se répartissent entre 6 « hotspots », ciblant les acides aminés R175, G245, R248, R249, R273 et R282, bien que des mutations de p53 ont été trouvées sur presque chaque codon de son DBD dans les cancers humains (Baugh et al., 2018) (Pitolli et al., 2019). (Cho et al., 1994) (Soussi and Lozano, 2005). Les mutations de contact peuvent préserver les propriétés thermodynamiques de la p53 WT mais altèrent sa capacité à se lier à l'ADN ou avec des protéines partenaires (Hagn et al., 2010). ...
Le système immunitaire joue un rôle important dans le contrôle et l'éradication du cancer. Des acteurs majeurs de la réponse immune antitumorale sont les cellules tueuses naturelles (ou cellules NK) et les lymphocytes T cytotoxiques (ou CTL), capable de reconnaitre et détruire des cellules tumorales par l’exocytose de perforine et de granzymes contenus dans leur granule cytotoxique. Il a été montré au sein du laboratoire l’implication de la protéine suppresseur de tumeur p53 dans cette voie apoptotique. Or, plus de 50% des tumeurs humaines présentent des mutations inactivatrices de p53 ce qui favorise le développement tumoral. De ce fait, l’inactivation fréquente de p53 dans les tumeurs humaines pourrait leur permettre d’échapper à la destruction par les CTL et les cellules NK.Dans ce contexte, mes travaux de thèse ont montré que la réactivation pharmacologique de la fonction de p53 sauvage dans des cellules tumorales exprimant une p53 mutée augmente leur susceptibilité à la lyse induite par les cellules NK grâce à l’induction d’un processus d’autophagie. De plus, j’ai cherché à déterminer le lien entre les mutations de p53 et l’expression à la surface des cellules tumorales de PD-L1 qui empêche l’activation optimale des cellules cytotoxiques et conduit à leur épuisement. Mes travaux actuels suggèrent que l’expression de p53 mutantes induits une surexpression de PD-L1 à la surface des cellules cancéreuses. Les mécanismes expliquant ce phénomène sont en cours d’études.
... Studies have shown that P53 can induce cell cycle arrest and promote DNA repair or induce apoptosis through multiple pathways (68). However, a large number of studies have shown that the mutation rate of P53 in human tumor cells is high, which are closely related to the occurrence of tumors (70)(71)(72). Importantly, our study reveals that MEG3 can affect the expression and function of telomere-related genes, such as telomerase reverse transcriptase TERT, telomerase RNA TERC, telomere repeat RNA sequence TERRA, telomere length maintenance protein POT1 Exo1, TRF2, SNM1B, CST/AAF, etc., However, knockdown of P53 abolishes the effect of MEG3 on the expression and function of these genes. ...
Background: MEG3 is down-regulated expression in several tumors and inhibits human tumorigenesis. But so far, the mechanism of MEG3 in tumorigenesis is still unclear.
Methods: Gene infection, cellular and molecular technologies and tumorigenesis test in vitro and in vivo were performed,respectively.
Results: our results indicate that MEG3 enhances the P53 expression by triggering the loading of P300 and RNA polymerase II onto its promoter regions dependent on HP1α. Moreover, MEG3 increases the methylation modification of histone H3 at the 27th lysine via P53. Furthermore, MEG3 inhibits the expression of TERT by increasing the H3K27me3 in TERT promoter regions. Thereby, MEG3 inhibits the activity of telomerase by reducing the binding of TERT to TERC. Furthermore, MEG3 also increases the expression of TERRA ,therefore, the interaction between TERC and TERT was competitively attenuated by increasing the interaction between TERRA and TERT, which inhibits the activity of telomerase in hLCSCs. Strikingly, MEG3 reduces the length of telomere by blocking the formation of complex maintaining telomere length(POT1-Exo1-TRF2-SNM1B) and decreasing the binding of the complex to telomere by increasing the interplay between P53 and HULC. Ultimately, MEG3 inhibits the growth of hLCSCs by reducing the activity of telomerase and attenuating telomeric repeat binding factor 2(TRF2).
Conclusions: Our results demonstrates MEG3 inhibits the occurrence of human liver cancer by blocking telomere and these findings provide an important insight into the prevention and treatment of human liver cancer.
... Studies have shown that P53 can induce cell cycle arrest and promote DNA repair or induce apoptosis through multiple pathways [64]. However, a large number of studies have shown that the mutation rate of P53 in human tumor cells is high, which are closely related to the occurrence of tumors [65][66][67]. Importantly, our study reveals that MEG3 can affect the expression and function of telomererelated genes, such as telomerase reverse transcriptase TERT, telomerase RNA TERC, telomere repeat RNA sequence TERRA, telomere length maintenance protein POT1 Exo1, TRF2, SNM1B, and CST/AAF. However, knockdown of P53 abolishes the effect of MEG3 on the expression and function of these genes. ...
Background
MEG3 downregulated the expression in several tumors and inhibits human tumorigenesis. But so far, the mechanism of MEG3 in tumorigenesis is still unclear.
Methods
In gene infection, cellular and molecular technologies and tumorigenesis test in vitro and in vivo were performed, respectively.
Results
Our results indicate that MEG3 enhances the P53 expression by triggering the loading of P300 and RNA polymerase II onto its promoter regions dependent on HP1α. Moreover, MEG3 increases the methylation modification of histone H3 at the 27th lysine via P53. Furthermore, MEG3 inhibits the expression of TERT by increasing the H3K27me3 in TERT promoter regions, thereby inhibiting the activity of telomerase by reducing the binding of TERT to TERC. Furthermore, MEG3 also increases the expression of TERRA; therefore, the interaction between TERC and TERT was competitively attenuated by increasing the interaction between TERRA and TERT, which inhibits the activity of telomerase in hLCSCs. Strikingly, MEG3 reduces the length of telomere by blocking the formation of complex maintaining telomere length (POT1-Exo1-TRF2-SNM1B) and decreasing the binding of the complex to telomere by increasing the interplay between P53 and HULC. Ultimately, MEG3 inhibits the growth of hLCSCs by reducing the activity of telomerase and attenuating telomeric repeat binding factor 2(TRF2).
Conclusions
Our results demonstrates MEG3 inhibits the occurrence of human liver cancer by blocking telomere, and these findings provide an important insight into the prevention and treatment of human liver cancer.
... However, the metabolism of effector T lymphocytes relies on more aerobic glycolysis, resulting in competition between effector T cells and cancer cells [54]. p53, a tumor suppressor protein, is frequently mutated in cancer cells [55,56], and local activation of p53 in the CME can overcome immune suppression and enhance anti-cancer activity [57]. ...
Modest levels of reactive oxygen species (ROS) are necessary for intracellular signaling, cell division, and enzyme activation. These ROS are later eliminated by the body’s antioxidant defense system. High amounts of ROS cause carcinogenesis by altering the signaling pathways associated with metabolism, proliferation, metastasis, and cell survival. Cancer cells exhibit enhanced ATP production and high ROS levels, which allow them to maintain elevated proliferation through metabolic reprograming. In order to prevent further ROS generation, cancer cells rely on more glycolysis to produce ATP and on the pentose phosphate pathway to provide NADPH. Pro-oxidant therapy can induce more ROS generation beyond the physiologic thresholds in cancer cells. Alternatively, antioxidant therapy can protect normal cells by activating cell survival signaling cascades, such as the nuclear factor erythroid 2-related factor 2 (Nrf2)-Kelch-like ECH-associated protein 1 (Keap1) pathway, in response to radio- and chemotherapeutic drugs. Nrf2 is a key regulator that protects cells from oxidative stress. Under normal conditions, Nrf2 is tightly bound to Keap1 and is ubiquitinated and degraded by the proteasome. However, under oxidative stress, or when treated with Nrf2 activators, Nrf2 is liberated from the Nrf2-Keap1 complex, translocated into the nucleus, and bound to the antioxidant response element in association with other factors. This cascade results in the expression of detoxifying enzymes, including NADH-quinone oxidoreductase 1 (NQO1) and heme oxygenase 1. NQO1 and cytochrome b5 reductase can neutralize ROS in the plasma membrane and induce a high NAD+/NADH ratio, which then activates SIRT1 and mitochondrial bioenergetics. NQO1 can also stabilize the tumor suppressor p53. Given their roles in cancer pathogenesis, redox homeostasis and the metabolic shift from glycolysis to oxidative phosphorylation (through activation of Nrf2 and NQO1) seem to be good targets for cancer therapy. Therefore, Nrf2 modulation and NQO1 stimulation could be important therapeutic targets for cancer prevention and treatment.
... The guardian of the genome, p53, is a key player in these cellular responses [57]. During tumor progression, pro-apoptotic mechanisms are lost mainly due to impaired functional p53 in more than 50% of human malignant tumors [58][59][60]. The ATM/ATR-activated p53 signals cell cycle arrest and DNA damage repair, thus promoting cell survival. ...
Although once considered as structural components of eukaryotic biological membranes, research in the past few decades hints at a major role of bioactive sphingolipids in mediating an array of physiological processes including cell survival, proliferation, inflammation, senescence, and death. A large body of evidence points to a fundamental role for the sphingolipid metabolic pathway in modulating the DNA damage response (DDR). The interplay between these two elements of cell signaling determines cell fate when cells are exposed to metabolic stress or ionizing radiation among other genotoxic agents. In this review, we aim to dissect the mediators of the DDR and how these interact with the different sphingolipid metabolites to mount various cellular responses.
... Modern science has no doubt about the enormous impact of mutated p53 alleles on carcinogenesis. Literature data suggest that almost 50% of all cancers have a mutated p53 gene [205]. There are several ways in which p53 can be targeted as a potential anti-cancer treatment, both in general and in hematological diseases. ...
Mitochondria are the main fascinating energetic source into the cells. Their number, shape, and dynamism are controlled by the cell's type and current behavior. The perturbation of the mitochondrial inward system via stress response and/or oncogenic insults could activate several trafficking molecular mechanisms with the intention to solve the problem. In this review, we aimed to clarify the crucial pathways in the mitochondrial system, dissecting the different metabolic defects, with a special emphasis on hematological malignancies. We investigated the pivotal role of mitochondria in the maintenance of hematopoietic stem cells (HSCs) and their main alterations that could induce malignant transformation, culminating in the generation of leukemic stem cells (LSCs). In addition, we presented an overview of LSCs mitochondrial dysregulated mechanisms in terms of (1) increasing in oxidative phosphorylation program (OXPHOS), as a crucial process for survival and self-renewal of LSCs,(2) low levels of reactive oxygen species (ROS), and (3) aberrant expression of B-cell lymphoma 2 (Bcl-2) with sustained mitophagy. Furthermore, these peculiarities may represent attractive new "hot spots" for mitochondrial-targeted therapy. Finally, we remark the potential of the LCS metabolic effectors to be exploited as novel therapeutic targets.
... It has also been reported that commonly inherited genetic polymorphisms in the p53 pathway increases the risk of a broad range of cancers more than polymorphisms in other pathways [31]. Previous studies have shown that, TP53 gene is very important for tumor suppression and apoptosis [32]. Moreover, homozygote Arg72 allele has 15 times more potency to induce apoptosis compared to its Pro72 counterpart [33].This is due to the fact that Arg72 allele can interact directly with proapoptotic BAK protein and thus makes it less penetrable for cancer susceptibility [34]. ...
Pharmacogenetic study of TP53 gene polymorphisms has not been conducted extensively in cervical cancer. The aim of this study was to assesses the TP53 codon 72 and codon 47 polymorphisms and their relation to cervical cancer risk in Bangladeshi women. 134 cervical cancer patients and 102 age matched healthy controls were included from two institutions in Bangladesh. Polymerase chain reaction-restriction fragment length polymorphism (PCR–RFLP) method was used for genotyping two TP53 single nucleotide polymorphisms (codon 72 and codon 47) in patients and controls. The results indicate that the TP53 Arg/Pro heterozygosity (adjusted OR 2.32, 95% CI 1.28–4.34, p = 0.01), Pro/Pro mutant homozygosity (adjusted OR 4.15, 95% CI 1.75–9.86, p = 0.001), along with the combined genotype (Arg/Pro + Pro/Pro) (adjusted OR 2.83, 95% CI 1.61–4.97, p < 0.001) significantly increases the risk of cervical cancer. Moreover, the cervical cancer patients with a first-degree relative cancer patient possesses 4.45 folds more risk (p = 0.019) of carrying a proline allele in codon 72 of the TP53 gene compared to those patients who do not have any first-degree relative with cancer. Finally, polymorphism in the codon 47 of the TP53 gene did not significantly increase the risk of cervical cancer in Bangladeshi women. To conclude, this is the first study to identify that polymorphism in the TP53 codon 72 significantly increases the risk of cervical cancer in a female population in Bangladesh.
... Depending on whether the corresponding residues are involved in DNA contact or structure maintenance, mutant proteins are categorized as contact (R273H, R248Q, and R248W) or conformational (R175H, G245S, R249S, and R282H) [27,28]. Contact mutants derive from missense mutations in residues responsible for direct contact with the DNA sequences forming p53 response elements in target gene promoters and have an intact native fold [29][30][31]. Conformational mutations result in the disruption of the p53 protein structure by decreasing the already low folding stability of the DBD, leading to its denaturation and often aggregation at body temperature [27]. Nevertheless, the distinction between these two mutation categories is somewhat arbitrary, as there are p53 mutants that, in principle, fit in both (e.g., R248Q) [27,32]. ...
p53 suppresses tumorigenesis by activating a plethora of effector pathways. While most of these operate primarily inside of cells to limit proliferation and survival of incipient cancer cells, many extend to the extracellular space. In particular, p53 controls expression and secretion of numerous extracellular factors that are either soluble or contained within extracellular vesicles such as exosomes. As part of the cellular secretome, they execute key roles in cell-cell communication and extracellular matrix remodeling. Mutations in the p53-encoding TP53 gene are the most frequent genetic alterations in cancer cells, and therefore, have profound impact on the composition of the tumor cell secretome. In this review, we discuss how the loss or dominant-negative inhibition of wild-type p53 in concert with a gain of neomorphic properties observed for many mutant p53 proteins, shapes a tumor cell secretome that creates a supportive microenvironment at the primary tumor site and primes niches in distant organs for future metastatic colonization.
... Residue Arg273 is essential for the DNA binding activity of the TP53 protein; changes at this position are known to disrupt TP53 tumor-suppressive activity [10]. TP53 gene mutations are the most common single genetic alteration in sporadic human epithelial ovarian cancer [4], almost all of them missense, largely occurring in the DNA binding domain, which may inactivate it and therefore reduce the ability to repress target proto-oncogenes [11]. Generally, the presence of a disruptive Tp53 mutation is associated with a poor prognosis in all types of cancers and diminished sensitivity to radiation and chemotherapy [12]. ...
Because of the profound heterogeneity of ovarian cancer at the clinical, cellular and molecular levels, herein we discuss the molecular findings at the protein and genetic levels seen in our patient. Immunohistochemistry showed a complete loss of phosphatase and tensin homolog, this observation was the reason behind prescribing the CDK4/6 inhibitor palbociclib. However, there was no response to treatment. Next-generation sequencing analysis was performed showing a nonsense mutation, p.R552X in retinoblastoma 1 ( RB1 ). This nonsense variation will possibly lead to a truncated protein lacking the domain responsible for interaction with E2F, an event that will induce cell cycle progression and, thus, be responsible for the chemo-resistance to palbociclib.
... In this regard, the central DNAbinding domain of p53 spans the most conserved region composed of a vast number of these missense mutations and among these the hot spot residues occur with unusually high frequency [2][3][4]. p53 missense mutations in the hot spot region can generally be classified as DNA contact (or class I) mutants, like R273H-p53, which normally make direct contact with target DNA sequences and conformational (or class II) mutants, like R175H-p53, which disrupt the structure of the p53 protein partially or completely, thus altering 2 Journal of Oncology its function [5,6]. R175H-P53 and R273H-P53, being the most frequently occurring GOF mutations in cancer cells, were observed to induce resistance to chemotherapeutic agents in multiple cancer cell types [7,8]. ...
Mutations in p53, especially gain of function (GOF) mutations, are highly frequent in lung cancers and are known to facilitate tumor aggressiveness. Yet, the links between mutant GOF-p53 and lung cancers are not well established. In the present study, we set to examine how we can better sensitize resistant GOF-p53 lung cancer cells through modulation of cellular protein degradation machineries, proteasome and autophagy. H1299 p53 null lung cancer cells were stably transfected with R273H mutant GOF-p53 or wild-type (wt) p53 or empty vectors. The presence of R273H-P53 conferred the cancer cells with drug resistance not only against the widely used chemotherapeutic agents like cisplatin (CDDP) or 5-flurouracil (5-FU) but also against potent alternative modes of therapy like proteasomal inhibition. Therefore, there is an urgent need for new strategies that can overcome GOF-p53 induced drug resistance and prolong patient survival following failure of standard therapies. We observed that the proteasomal inhibitor, peptide aldehyde N-acetyl-leu-leu-norleucinal (commonly termed as ALLN), caused an activation of cellular homeostatic machinery, autophagy in R273H-P53 cells. Interestingly, inhibition of autophagy by chloroquine (CQ) alone or in combination with ALLN failed to induce enhanced cell death in the R273H-P53 cells; however, in contrast, an activation of autophagy by serum starvation or rapamycin increased sensitivity of cells to ALLN-induced cytotoxicity. An activated autophagy was associated with increased ROS and ERK signaling and an inhibition of either ROS or ERK signaling resulted in reduced cytotoxicity. Furthermore, inhibition of GOF-p53 was found to enhance autophagy resulting in increased cell death. Our findings provide novel insights pertaining to mechanisms by which a GOF-p53 harboring lung cancer cell is better sensitized, which can lead to the development of advanced therapy against resistant lung cancer cells.
... TP53 gene which is tumor suppressor gene regulates DNA repair mechanism and apoptosis. The TP53 mutation is observed in nearly 50% CRCs, with the most frequently observed mutations between exons 5 and 8 at codons 175, 213, 245, 248, 273, and 282 [25,26]. Currently, molecular-based targeted therapy is a standard approach in selected patients such as those with unresected metastatic CRC disease being eligible for inhibitors of VEGF and EGFR. ...
Background: Colorectal cancer (CRC) has been identified as the third most common cancer worldwide. Gene mutation and defective cell regulation are important processes in the development of CRC. Assessment of genetic mutations is an essential element in the modern era of personalized cancer treatment.
... The most commonly mutated areas in the DBD are loop-sheet-binding helices (LSHs) L2 and L3. Various mutations have different consequences for the function of the p53 protein 12,13 . Some mutations are associated with a loss of function and others with a gain of function. ...
Diffuse large B-cell lymphoma (DLBCL) is an aggressive lymphoma with diverse outcomes. Concurrent translocation of MYC and BCL-2 and/or BCL-6, and concurrent immunohistochemical (IHC) high expression of MYC and BCL-2, have been linked to unfavorable treatment responses. TP53-mutated DLBCL has also been linked to worse outcome. Our aim was to evaluate the aforementioned issues in a cohort of 155 patients uniformly treated with R-CHOP-like therapies. We performed direct sequencing of TP53 exons 5, 6, 7 and 8 as well as fluorescence in-situ hybridization (FISH) of MYC, BCL-2 and BCL-6, and IHC of MYC, BCL-2 and BCL-6. In multivariate analysis, TP53 mutations in L3 and loop-sheet helix (LSH) associated with a risk ratio (RR) of disease-specific survival (DSS) of 8.779 (p = 0.022) and a RR of disease-free survival (DFS) of 10.498 (p = 0.011). In IHC analysis BCL-2 overexpression was associated with inferior DFS (p = 0.002) and DSS (p = 0.002). DLBCL with BCL-2 and MYC overexpression conferred inferior survival in all patients (DSS, p = 0.038 and DFS, p = 0.011) and in patients with non-GC phenotype (DSS (p = 0.013) and DFS (p = 0.010). Our results imply that in DLBCL, the location of TP53 mutations and IHC analysis of BCL-2 and MYC might have a role in the assessment of prognosis.
... The c.215G>C, p. (Arg72Pro) (rs1042522) SNP is among the most extensively studied polymorphisms found in the coding region of TP53. It is located in codon 72 of exon 4, in a proline-rich domain, which is known to be important for the induction of apoptosis by p53 [12,14]. ...
Background:
The purpose of our case control study is to explore the potential association of tumor protein 53 (TP53) c.215G>C, p. (Arg72Pro) polymorphism (rs1042522) with the risk of breast cancer (BC) development in the Moroccan population.
Methods and results:
The study population consisted of 125 female patients with confirmed BC and 126 healthy controls. DNA samples were genotyped by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism assay method using BstUI restriction enzyme. We showed that the homozygous genotype of TP53 72Pro variant was significantly associated with increased BC risk (OR 2.2, 95% CI 1.07-4.54, p = 0.03). The dominant and additive models of TP53 Pro allele were also correlated to the risk of BC (OR 2.13, 95% CI 1.07-4.23, p = 0.02 and OR 1.49, 95% CI 1.03-2.16, p = 0.03, respectively). Furthermore, the TP53 Arg72 variant was associated with protection against BC, either in the homozygous genotype, the dominant or the additive models (OR 0.45, 95% CI 0.22-0.93, p = 0.03; OR 0.46, 95% CI 0.23-0.92, p = 0.029 and OR 0.67, 95% CI 0.46-0.97, p = 0.03, respectively).
Conclusion:
Our results suggest that TP53 c.215G>C, p. (Arg72Pro) polymorphism may be considered as a genetic marker for predisposition to BC in Moroccan population.
... Despite being genetically heterozygous at the nucleotides encoding L332, CTB-m2 cells expressed almost only the mutated L332Q p53 mRNA suggesting that in these cells the wild-type p53 allele is inactive. The expression of p53 mutants has been shown to be heterogenous and gene methylation has also been observed in human cancers (25,26). CTB-m2 cells may therefore have a unique expression status among p53 mutant alleles. ...
Mutations in the p53 gene are associated with more than half of all human cancers. These mutations often cause a disruption of the tumor-suppressor function of p53 and induce genomic instabilities. Wild‑type p53 requires tetramerization to function as an initiator of cell cycle arrest and apoptosis. Although alterations in p53 tetramerization caused by mutation have been well studied, there are few cell lines containing an endogenous mutation in the tetramerization domain of p53. Here, we report the discovery of a canine mammary gland tumor cell line CTB‑m2, which contains the Leu332Gln (L332Q) mutation corresponding to Leu344 in the tetramerization domain of human p53. Although CTB‑m2 cells are genetically heterozygous for the Leu332Gln mutation, the mutant mRNA was almost exclusively expressed. CTB‑m2 cells showed enhanced cell proliferation compared to wild‑type p53-expressing CTB‑m cells of the same lineage. A p53 tetramerization reporter assay showed that the ability of the p53 mutant to form tetramers was significantly lower than that of wild‑type p53. An immunoblot analysis of cross-linked p53 oligomerized forms demonstrated that the L332Q mutant lacked the ability to form tetramers but retained the ability to form dimers. These data suggest that the p53 mutant cell line CTB‑m2 could be a useful tool for analyzing the precise tetramerization mechanisms of p53 and verifying the effects of therapeutic agents against tumors expressing p53 mutants that lack the ability to tetramerize.
... Plusieurs facteurs semblent intervenir dans ce choix: l'importance des lésions de l'ADN, le niveau d'expression de P53 induit, le type cellulaire ou l'altération d'autres oncogènes. Après exposition des cellules à un stress génotoxique le niveau d'expression de P53 augmente, et il est accompagné de son activation suite à des modifications posttraductionnelles. Cette induction peut conduire soit à l'arrêt du cycle cellulaire en phase G1 afin de donner le temps à l'ADN cellulaire d'être réparé avant de rentrer en phase S soit à l'apoptose dans le but d'éliminer les cellules contenant une altération génétique.Figure 5: Le pourcentage des mutations de p53 par type d'organe.Contrairement aux autres gènes suppresseurs de tumeurs qui sont inactivés par troncation ou délétion, plus de 80% des altérations de P53 sont dues à des mutations faux-sens qui stabilisent le niveau d'expression de p53(Soussi and Lozano 2005). ...
L'inactivation des gènes p53 et pten présente un facteur de mauvais pronostic et de résistance à différents traitements anticancéreux incluant les thérapies ciblées. Le cétuximab (Erbitux®) est un anticorps monoclonal chimérique dirigé contre le domaine extracellulaire du récepteur à l'EGF. Son mécanisme d'action est fortement lié à la fonctionnalité des voies de signalisation de PI3KJAKT et des MAPK. Nous avons évalué l'influence de la réintroduction des gènes p53 et pten par vectorisation non viral à base de polyéthylènimine, couplée ou non à l'internalisation photochimique sur l'induction de l'apoptose et l'inhibition de la croissance cellulaire dans des cellules P53 et pten mutés d'une part, et sur l'impact de ce transfert de gène sur la fonctionnalité des voies de signalisation impliquées dans la réponse cellulaire au cétuximab. Nous avons montré que la transfection des gènes suppresseurs de tumeurs p53 et pten par polyétylènimine couplé à la PCI rétablit l'expression de P53 et de PTEN et permet de restaurer l'inhibition de croissance et l'induction de l'apoptose. Associée au traitement par cétuximab La restauration de PTEN et P53 réprime l'activation constitutive des voies de signalisation PI3K et MAPK et potentialise l'inhibition de la croissance et l'induction de l'apoptose induites par le cétuximab. Ces résultats montrent que l'inactivation de p53 et pten pourrait être prédictive de la résistance au cétuximab à travers l'activation constitutive de la voie de signalisation PI3/AKT et confirme que la détennination de la fonctionnalité des voies de signalisation des rtiveaux d'expression des phosphoprotéines de signalisation permettrait de prédire la réponse au cétuximab et de proposer des options thérapeutiques originales.
In 2021, the World Health Organization released the fifth edition of the central nervous system (CNS) tumor classification. This classification uses histopathology and molecular pathogenesis to group tumors into more biologically and molecularly defined entities. The prognosis of brain cancer, particularly malignant tumors, has remained poor worldwide, approximately 308,102 new cases of brain and other CNS tumors were diagnosed in the year 2020, with an estimated 251,329 deaths. The cost and time-consuming nature of studies to find new anticancer agents makes it necessary to have well-designed studies. In the present study, the pathways that can be targeted for drug development are discussed in detail. Some of the important cellular origins, signaling, and pathways involved in the efficacy of bioactive molecules against CNS tumorigenesis or progression, as well as prognosis and common approaches for treatment of different types of brain tumors, are reviewed. Moreover, different study tools, including cell lines, in vitro, in vivo, and clinical trial challenges, are discussed. In addition, in this article, natural products as one of the most important sources for finding new chemotherapeutics were reviewed and over 700 reported molecules with efficacy against CNS cancer cells are gathered and Drug Dev Res. 2024;85:e22180. wileyonlinelibrary.com/journal/ddr Abbreviations: 4EBP1, eukaryotic initiation factor binding protein 1; AKT, protein kinase B; ATM, ataxia telangiectasia mutated; BBB, blood brain barrier; CAT, catalytic subunit; CDK, cyclin-dependent kinase; CDKs, cyclin-dependent kinases; CHOP, C/EBP HOMOLOGOUS PROTein; CK2, casein kinase 2; CNS, central nervous system; COX, cyclooxygenase; CSF, cerebrospinal fluid; cyt c, Cytochrome c; EC 50 , half maximal effective concentration; ED 50 , median effective dose, which is the dose that produces the effect in 50% of the population that take that dose; EGF, epidermal growth factor; EGFR, epidermal growth factor receptor; eIF4E, eukaryotic initiation factor 4E; ER, endoplasmic reticulum; ERK, extracellular-signal-regulated kinase; FGFR, fibroblast growth factor receptor; FPP, farnesyl pyrophosphate; FTase, farnesyltransferase; GB, glioblastoma; GBM, glioblastoma multiforme; GDP, guanosine diphosphate; GEFs, guanine nucleotide exchange factors; GFAP, glial fibrillary acidic protein; GGTase-I, geranylgeranyltransferase type I; GI50, concentration of any compound required for 50% growth inhibition of cells; Grb2, growth factor receptor-bound protein 2; GSK, glycogen synthase kinase; GTP, guanosine triphosphate; HAAE-2, human abdominal aorta endothelial cells; HDM2, human and/or murine double minute-2 protein; HIF-1α, hypoxia-inducible factor 1 alpha; HMGB1, high mobility group box 1 protein; IC 50 , half maximal inhibitory concentration; JNK, c-Jun N-terminal kinase; LOX, lipoxygenase; LPT, long-term potentiation; LRP, lipoprotein related proteins; LXRs, liver X receptors; mAB, monoclonal antibodies; MAPK, mitogen-activated protein kinase; MAPK, mitogen-activated protein kinase; MAST, microtubule-associated serine-threonine kinase; MDM2, p53-Mouse double minute 2; MEKis, MEK inhibitors; MMP, matrix metalloproteinase; mTORC1, mammalian target of rapamycin complex 1; MVA, mevalonate; MW, molecular weight; NAPA, Napabucasin; NF1, Neurofibromin 1; NIH, National Institutes of Health; NQO1, NAD(P)H Quinone Dehydrogenase 1; NSCs, neural stem cells; PA, pilocytic astrocytoma; PDGFR, platelet-derived growth factor receptor; PDGFRs, platelet-derived growth factor receptors; PERK, protein kinase RNA-like endoplasmic reticulum kinase (PERK); PFS, progression-free survival; PI3K, phosphatidylinositol-4,5-bisphosphate 3-kinase; PI3K, phosphatidylinositol-4,5-bisphosphate 3-kinase; PI3P, intracellular phosphatidylinositol-3,4,5-trisphosphate; PI(3,4,5)P3 or PIP3, phosphatidylinositol (3,4,5)-trisphosphate; PI(4,5)P2 or PIP2, phosphatidylinositol (4,5)-bisphosphate; pRB, retinoblastoma protein; PTEN, phosphatase and tensin homolog deleted on chromosome 10; PTM, posttranslational modification; RAGE, receptor for advanced glycation end products; RECK, reversion-inducing-cysteine-rich protein with kazal motifs; ROCK, RhoA-associated kinase; ROS, reactive oxygen species; RTKs, receptor tyrosine kinases; SAPKs, stress-activated protein kinases; SC, subcutaneous; Sp1, specificity protein 1; TAD, transactivation domain; TGI, concentration eliciting total growth inhibition; TKIs, tyrosine kinase inhibitors; TP53, tumor protein p53; TUNEL, terminal uridine deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling; VEGFRs, vascular endothelial growth factor receptors; WHO, World Health Organization. Ardalan Pasdaran and Azadeh Hamedi gathered data. All authors contributed in writing and revising the manuscript.
In 2021, the World Health Organization released the fifth edition of the central nervous system (CNS) tumor classification. This classification uses histopathology and molecular pathogenesis to group tumors into more biologically and molecularly defined entities. The prognosis of brain cancer, particularly malignant tumors, has remained poor worldwide, approximately 308,102 new cases of brain and other CNS tumors were diagnosed in the year 2020, with an estimated 251,329 deaths. The cost and time‐consuming nature of studies to find new anticancer agents makes it necessary to have well‐designed studies. In the present study, the pathways that can be targeted for drug development are discussed in detail. Some of the important cellular origins, signaling, and pathways involved in the efficacy of bioactive molecules against CNS tumorigenesis or progression, as well as prognosis and common approaches for treatment of different types of brain tumors, are reviewed. Moreover, different study tools, including cell lines, in vitro, in vivo, and clinical trial challenges, are discussed. In addition, in this article, natural products as one of the most important sources for finding new chemotherapeutics were reviewed and over 700 reported molecules with efficacy against CNS cancer cells are gathered and classified according to their structure. Based on the clinical trials that have been registered, very few of these natural or semi‐synthetic derivatives have been studied in humans. The review can help researchers understand the involved mechanisms and design new goal‐oriented studies for drug development against CNS malignancies.
Among the tumor suppressor genes, TP53 is the most frequently mutated in human cancers, and most mutations are missense mutations causing production of mutant p53 (mutp53) proteins. TP53 mutations not only results in loss of function (LOH) as a transcription factor and a tumor suppressor, but also gain wild-type p53 (WTp53)-independent oncogenic functions that enhance cancer metastasis and progression (Yamamoto and Iwakuma, 2018; Zhang et al., 2022). TP53 has extensively been studied as a therapeutic target as well as for drug development and therapies, however with limited success. Achieving targeted therapies for restoration of WTp53 function and depletion or repair of mutant p53 (mutp53) will have far reaching implication in cancer treatment and therapies. This review briefly discusses the role of p53 mutation in cancer and the therapeutic potential of restoring WTp53 through the advances in mRNA nanomedicine.
Mutations in the p53 gene are common and occur in over 50% of all cancers, as it is involved in DNA damage repair, cell cycle regulation and apoptosis. Moreover, the p53 gene is mutated in 70% of colon cancers. Therefore, the development of drugs to combat this mutation requires urgent attention. With this in mind, in silico drug design approaches were applied on quinoline derivatives with anticancer activity. In 3D-QSAR study, steric, electrostatic, hydrophobic and H-bond acceptor fields (SEHA) play an important role in prediction and design of new colon cancer compounds. Indeed, the two best CoMSIA/SEHA models with (Q² = 0.737, R² = 0.914, Rpred2 = 0.720) and (Q² = 0.738, R² = 0.919, Rpred2= 0.739) show good prediction of human colon carcinoma HCT 116 (p53+/+) and (p53−/−) activities, respectively. Furthermore, the predictive ability and robustness of these models were tested by several validation methods. Molecular docking analyses reveal crucial interactions with the active sites of the p53 protein in both wild type and mutant. Based on these theoretical studies, we designed 10 new compounds with good anticancer activity potential, which were evaluated using ADMET properties. Molecular dynamics simulations were performed to confirm the detailed binding mode of the docking results. Finally, the MM-GBSA based on molecular dynamics simulation confirmed that the designed compounds were able to form stable hydrogen bonding interactions with the crucial residues, which are essential to overcome the p53 mutation in colon cancer.
Communicated by Ramaswamy H. Sarma
Purpose:
Cataract is a leading visual disease characterized by enhanced oxidative stress and increased apoptosis of human lens epithelial cells (HLECs). TRIM3 is a tumor suppressor in many cancers. However, its role in cataract remains unknown. In this study, we aimed to explore the role of TRIM3 in H2O2-injured HLECs and the underlying mechanisms involved.
Methods:
HLECs were treated with different H2O2 concentrations to induce apoptosis. A lentivirus was designed to overexpress TRIM3 and p53, and TRIM3 knockdown was prepared. A P53 inhibitor, PFTα, was used to knockdown p53. Cell viability and apoptosis were detected by CCK-8 and flow cytometric analyses, respectively. TRIM3, p53, Bcl2, and Bax expression levels were determined by qRT-qPCR and western blotting.
Results:
It was found that H2O2-treated HLECs had markedly decreased cell viability and TRIM3 expression. TRIM3 overexpression attenuated the H2O2-induced HLEC apoptosis, while TRIM3 knockdown promoted it. P53, a downstream target of TRIM3, was found to be negatively regulated by TRIM3 via ubiquitination in HLECs. Furthermore, p53 overexpression abolished the effect of TRIM3 overexpression on H2O2-induced HLEC apoptosis, while PFTα alleviated the TRIM3 knockdown-mediated HLEC apoptosis.
Conclusion:
This study demonstrates that TRIM3 inhibited the H2O2-induced apoptosis of HLECs by decreasing p53 via ubiquitination.
P53 is known as the most critical tumor suppressor and is often referred to as the guardian of our genome. More than 40 years after its discovery, we are still struggling to understand all molecular details on how this transcription factor prevents oncogenesis or how to leverage current knowledge about its function to improve cancer treatment. Multiple cues, including DNA-damage or mitotic errors, can lead to the stabilization and nuclear translocation of p53, initiating the expression of multiple target genes. These transcriptional programs may be cell-type- and stimulus-specific, as is their outcome that ultimately imposes a barrier to cellular transformation. Cell cycle arrest and cell death are two well-studied consequences of p53 activation, but, while being considered critical, they do not fully explain the consequences of p53 loss-of-function phenotypes in cancer. Here, we discuss how mitotic errors alert the p53 network and give an overview of multiple ways that p53 can trigger cell death. We argue that a comparative analysis of different types of p53 responses, elicited by different triggers in a time-resolved manner in well-defined model systems, is critical to understand the cell-type-specific cell fate induced by p53 upon its activation in order to resolve the remaining mystery of its tumor-suppressive function.
A variety of biological processes are regulated by posttranslational modifications. Posttranslational modifications including phosphorylation, ubiquitination, glycosylation, and proteolytic cleavage, control diverse physiological functions in the gastrointestinal tract. Therefore, a better understanding of their implications in intestinal diseases, including inflammatory bowel disease, irritable bowel syndrome, celiac disease, and colorectal cancer would provide a basis for the identification of novel biomarkers as well as attractive therapeutic targets. Posttranslational modifications can be common denominators, as well as distinct biomarkers, characterizing pathological differences of various intestinal diseases. This review provides experimental evidence that identifies changes in posttranslational modifications from patient samples, primary cells, or cell lines in intestinal disorders, and a summary of carefully selected information on the use of pharmacological modulators of protein modifications as therapeutic options.
Majority of cancers harbor alterations of the tumor suppressor TP53 . However, childhood cancers, including unfavorable neuroblastoma, often lack TP53 mutations despite frequent loss of p53 function, suggesting alternative p53 inactivating mechanisms.
Here we show that p53-regulating PPM1D at chromosome 17q22.3 is linked to aggressive tumors and poor prognosis in neuroblastoma. We identified that WIP1-phosphatase encoded by PPM1D , is activated by frequent segmental 17q-gain further accumulated during clonal evolution, gene-amplifications, gene-fusions or gain-of-function somatic and germline mutations. Pharmacological and genetic manipulation established WIP1 as a druggable target in neuroblastoma. Genome-scale CRISPR-Cas9 screening demonstrated PPM1D genetic dependency in TP53 wild-type neuroblastoma cell lines, and shRNA PPM1D knockdown significantly delayed in vivo tumor formation. Establishing a transgenic mouse model overexpressing PPM1D showed that these mice develop cancers phenotypically and genetically similar to tumors arising in mice with dysfunctional p53 when subjected to low-dose irradiation. Tumors include T-cell lymphomas harboring Notch1 -mutations, Pten -deletions and p53-accumulation, adenocarcinomas and PHOX2B- expressing neuroblastomas establishing PPM1D as a bona fide oncogene in wtTP53 cancer and childhood neuroblastoma. Pharmacological inhibition of WIP1 suppressed the growth of neural tumors in nude mice proposing WIP1 as a therapeutic target in neural childhood tumors.
Trotz der Fortschritte in der Therapie des Multiplen Myeloms und des stetig wachsenden Arsenals effektiver anti-MM-Medikamente muss ein Teil der Patienten mit bestimmten zytogenetischen Veränderungen der Tumorzellen nach wie vor der Hochrisiko-Gruppe zugeordnet werden und hat eine Lebens-erwartung von nur wenigen Jahren. Einer der ungünstigsten prognostischen Marker ist die Inaktivierung des Tumorsuppressorgens TP53 durch Mutationen des Gens oder Deletionen des kurzen Arms von Chromosom 17, del(17p). Diese wird häufig mit einer Chemoresistenz der entarteten Plasmazellen in Verbindung gebracht. In der vorliegenden Arbeit gelang es mittels des CRISPR/Cas9-Systems TP53-Läsionen zu erzeugen und isogene Klone der TP53wt/wt Zelllinie AMO-1 zu generieren. Diese wurden anhand der Sequenzanalysen von beiden TP53-Allelen den Gruppen der biallelisch TP53-inaktivierten, der monoallelisch TP53-inaktivierten und der TP53wt/wt Klone zugeordnet. Das gruppenspezifische Verhalten der Klone aller drei Gruppen hinsichtlich deren Expression von p53, p21 und Mdm2 unterstrich die Validität des etablierten Zelllinienmodells zur Untersuchung der Bedeutung von TP53-Läsionen beim Multiplen Myelom. Neben einer kompletten Ausschaltung durch biallelische TP53-Inaktivierung zeigten die Ergebnisse der vorliegenden Arbeit auch eine Haploinsuffizienz des p53-Systems. Diese äußerte sich in einer Abschwächung der Nutlin-3A-abhängigen p53-, p21- und Mdm2-Induktion bereits nach Inaktivierung eines TP53-Allels durch Frameshift-Mutation. Korrelierend zu dem Proteinexpressions¬muster konnte eine zunehmende Resistenzentwicklung der Klone je nach Grad der TP53-Inaktivierung (mono- bzw. biallelisch) gegen Nutlin 3A sowie genotoxische Substanzen nachgewiesen werden, während die Sensibilität der MM-Zellen gegen Proteasominhibitoren unbeeinträchtigt blieb. Einschränkungen hinsichtlich der Übertragbarkeit der Ergebnisse der vorliegenden Arbeit auf das Multiple Myelom im Allgemeinen bestehen in dem Umstand, dass die beschriebenen Beobachtungen lediglich an einer einzigen MM-Zelllinie gemacht werden konnten. Dies ist durch die geringe Auswahl an TP53wt/wt MM-Zelllinien, die zudem noch oft eine schlechte Transfektabilität und niedrige Zellteilungsrate nach Einzelzellselektion aufweisen, bedingt. Die an der Zelllinie AMO-1 gemachten Beobachtungen stehen in Einklang mit der in klinischen Studien festgestellten Verkürzung des progressionsfreien- (PFS) und Gesamt-Überlebens (OS) bei MM-Patienten mit TP53-Alterationen. Die zunehmende Chemoresistenz der malignen Plasmazellen nach mono- bzw. biallelischer TP53-Inaktivierung kann als Grund für die Akkumulation entsprechender Klone im Rezidiv und in fortgeschrittenen Krankheitsstadien des MM angesehen werden. Mittels möglichst umfassender Erfassung des genauen TP53-Läsions-Status in zukünftigen klinischen Studien zu multiplen Zeitpunkten des Krankheitsverlaufs könnte der Einfluss verschiedener, in der Therapie des MM zum Einsatz kommender Substanzen auf die Selektion bzw. die Unterdrückung besonders virulenter Subklone mit TP53-Läsionen untersucht werden.
Background and Aims
p53 mutations occur frequently in human hepatocellular carcinoma (HCC). Activation of the mammalian target of rapamycin (mTOR) pathway is also associated with HCC. However, it is still unknown whether these changes together initiate HCC and can be targeted as a potential therapeutic strategy.
Methods
We generated mouse models in which mTOR was hyperactivated by loss of tuberous sclerosis complex 1 (Tsc1) with or without p53 haplodeficiency. Primary cells were isolated from mouse livers. Oncogenic signaling was assessed in vitro and in vivo, with or without targeted inhibition of a single molecule or multiple molecules. Transcriptional profiling was used to identify biomarkers predictive of HCC. Human HCC materials were used to corroborate the findings from mouse models.
Results
p53 haploinsufficiency facilitates mTOR signaling via the Pten/PI3k/Akt axis, promoting HCC tumorigenesis and lung metastasis. Inhibition of PI3K/Akt reduced mTOR activity, which effectively enhanced the anticancer effort of an mTOR inhibitor. Abcc4 was found to be responsible for p53 haploinsufficiency- and Tsc1 loss-driven HCC tumorigenesis. Moreover, in clinical HCC samples, Abcc4 specifically identified an aggressive subtype. The mTOR inhibitor rapamycin significantly reduced hepatocarcinogenesis triggered by Tsc1 loss and p53 haploinsufficiency in vivo, as well as the biomarker Abcc4.
Conclusions
Our data advance the current understanding of the activation of the Pten/PI3K/Akt/mTOR axis and its downstream target Abcc4 in hepatocarcinogenesis driven by p53 reduction and Tsc1 loss. Targeting mTOR, an unexpected vulnerability in p53 (haplo)deficiency HCC, can be exploited therapeutically to treat Abcc-4-positive HCC patients.
Small molecule modulation of protein-protein interactions is a very promising but also challenging area in drug discovery. The tumor suppressor protein p53 is one of the most frequently altered proteins in human cancers, making it an attractive target in oncology. 14-3-3 proteins have been shown to bind to and positively regulate p53 activity by protecting it from MDM2-dependent degradation or activating its DNA binding affinity. Protein-protein interactions can be modulated by inhibiting or stabilizing specific interactions by small molecules. Whereas inhibition has been widely explored by pharmaceutical industry and academia, the opposite strategy of stabilizing protein-protein interactions still remains relatively underexploited. This is rather interesting considering the number of natural compounds like rapamycin, forskolin and fusicoccin that exert their activity by stabilizing specific protein-protein interactions. In this review, we give an overview of 14-3-3 interactions with p53, explain isoform specific stabilization of the tumor suppressor protein, explore the approach of stabilizing the 14-3-3σ - p53 complex and summarize some promising small-molecules inhibiting the p53 - MDM2 protein-protein interaction.
Background. TP53 gene mutations impede cell apoptosis and lead to additional oncogenic events contributing to tumor progression.
Aim. To assess TP53 gene mutation rate in patients with high-grade B-cell lymphoma double-hit (HGBCL DH) and not otherwise specified (HGBL NOS); to analyse its relationship to disease prognosis.
Materials & Methods. Retrospective materials from medical records of 10 HGBL DH and 26 HGBL NOS patients were analyzed. Median follow-up was 26.5 months (range 0.6–160.9 months). Selection was based on the presence of available biological materials (paraffin blocks) for Sanger sequencing of TP53 gene from exon 5 to exon 8 (encoding DNA-binding domain of TP53 gene). FISH analysis of the tumor was performed in all patients to identify translocations involving c–MYC/8q24, BCL2/18q21, and BCL6/3q27 gene locus. To analyze differences between groups χ2 and Mann-Whitney tests were applied. Univariate event analysis (Kaplan-Meier and log-rank tests) and Cox regression analysis were used to assess the influence of molecular markers on the disease prognosis.
Results. TP53 gene mutations in lymphoma cells were found in 13 (36 %) out of 36 patients, 10/ (77 %) out of 13 mutations were pathogenic. In 8 out of 10 patients with TP53 mutations c–MYC/8q24 gene translocation was identified. Groups with wild (TP53-WT) and mutant (TP53-MUT) types of TP53 gene were similar in terms of main clinical characteristics. Patients with TP53-MUT in tumor cells showed worse 3-year overall survival (OS) compared with the group without TP53-MUT (30 % vs. 73 %; p = 0.026) as well as higher probability of disease progression in the period of 3 years (66 % vs. 15 %; p = 0.004). In multivariate analysis significant OS predictor proved to be the presence of TP53 mutation (p = 0.006). Relapse/progression probability was higher in combined cases of TP53 mutation and translocation involving c–MYC gene locus (p = 0.0003).
Conclusion. Translocation involving c–MYC gene along with TP53 gene mutation in tumor cells can serve as a criterion for dividing HGBL DH and HGBL NOS patients into different lymphoma relapse/progression risk groups.
Exchange of macromolecules between the nucleo- and cytoplasm is provided by the nuclear transport system (NTS). Karyopherins represent essential components of NTS) by serving as nuclear transport receptors/adaptor proteins. Dysregulation of karyopherins in (hepato ) carcinogenesis, including the pivotal nuclear import factor karyopherin-α2 (KPNA2), has been previously reported. However, the functional and regulatory role of KPNA2 in hepatocellular carcinoma (HCC) remains incompletely understood. To further characterize KPNA2 in this context, mass spectrometry-based proteomics was combined with functional/mechanistic cell-based approaches and data derived from murine and human HCC samples. Quantitative mass spectrometry upon siRNA-mediated KPNA2 knockdown in HCC cells revealed the microtubule-related oncoprotein stathmin among the most downregulated proteins. KPNA2 depletion resulted in impaired colony formation and tumor cell migration in HCC cells, which could be recapitulated by direct knockdown of stathmin. Having identified stathmin as functional relevant “downstream” target of KPNA2 the underlying molecular mechanism of KPNA2-dependent stathmin regulation was dissected. Out of several candidates the transcription factors E2F1 and TFDP1 were identified as transport substrates of KPNA2 and to be retained in the cytoplasm upon KPNA2 ablation followed by reduced STMN1 expression. Finally, significant correlations of STMN1 with E2F1/TFDP1 and KPNA2 expression were found based on data derived from murine HCC models and human HCC cohorts with high KPNA2 and STMN1 expression being associated with poorer patient outcome. Taken together, these data suggest that KPNA2 regulates STMN1 by mediating nuclear import of E2F1/TFDP1 and thereby provide a functionally relevant link between the NTS and microtubule-interacting proteins in HCC. Though further studies are required, interfering with nuclear import factors such as KPNA2 could represent a promising therapeutic approach in liver cancer.
Transcriptional activation of p53-regulated genes is initiated by sequence-specific DNA binding of p53 to target binding sites. Regulation of sequence-specific DNA binding is complex and occurs at various levels. We demonstrate that DNA topology is an important parameter for regulating the selective and highly specific interaction of p53 with its target binding sites. Specific binding of wild-type p53 is greatly enhanced when cognate binding sites are present in a non-linear stem- loop conformation. The C-terminal domain plays a key role in regulating the specific interactions of p53 with target binding sites in a DNA conformation-dependent manner. The C-terminal domain is required for binding to target sites in a non-linear DNA conformation in contrast to the strong inhibitory effects of the C terminus on p53 interaction with linear DNA. We propose that selective binding of p53 to various promoters may be determined by the DNA conformation within p53 cognate sites.
Most of the oncogenic mutations in the tumor suppressor p53 map to its DNA-binding (core) domain. It is thus a potential target in cancer therapy for rescue by drugs. To begin to understand how mutation inactivates p53 and hence to provide a structural basis for drug design, we have compared structures of wild-type and mutant p53 core domains in solution by NMR spectroscopy. Structural changes introduced by five hot-spot mutations (V143A, G245S, R248Q, R249S, and R273H) were monitored by chemical-shift changes. Only localized changes are observed for G245S, R248Q, R249S, and R273H, suggesting that the overall tertiary folds of these mutant proteins are similar to that of wild type. Structural changes in R273H are found mainly in the loop–sheet–helix motif and the loop L3 of the core domain. Mutations in L3 (G245S, R248Q, and R249S) introduce structural changes in the loop L2 and L3 as well as terminal residues of strands 4, 9, and 10. It is noteworthy that R248Q, which is often regarded as a contact mutant that affects only interactions with DNA, introduces structural changes as extensive as the other loop L3 mutations (G245S and R249S). These changes suggest that R248Q is also a structural mutant that perturbs the structure of loop L2–L3 regions of the p53 core domain. In contrast to other mutants, replacement of the core residue valine 143 to alanine causes chemical-shift changes in almost all residues in the β-sandwich and the DNA-binding surface. Long-range effects of V143A mutation may affect the specificity of DNA binding.
Drug resistance in human cancer is associated with overexpression of the multidrug resistance (MDR1) gene, which confers cross-resistance
to hydrophobic natural product cytotoxic drugs. Expression of the MDR1 gene can occur de novo in human cancers in the absence
of drug treatment. The promoter of the human MDR1 gene was shown to be a target for the c-Ha-Ras-1 oncogene and the p53 tumor
suppressor gene products, both of which are associated with tumor progression. The stimulatory effect of c-Ha-Ras-1 was not
specific for the MDR1 promoter alone, whereas a mutant p53 specifically stimulated the MDR1 promoter and wild-type p53 exerted
specific repression. These results imply that the MDR1 gene could be activated during tumor progression associated with mutations
in Ras and p53.
Mutations in the p53 tumour-suppressor gene are the most frequently observed genetic lesions in human cancers. To investigate the role of the p53 gene in mammalian development and tumorigenesis, a null mutation was introduced into the gene by homologous recombination in murine embryonic stem cells. Mice homozygous for the null allele appear normal but are prone to the spontaneous development of a variety of neoplasms by 6 months of age. These observations indicate that a normal p53 gene is dispensable for embryonic development, that its absence predisposes the animal to neoplastic disease, and that an oncogenic mutant form of p53 is not obligatory for the genesis of many types of tumours.
Transfection of a cloned p53 gene into a murine bladder carcinoma cell with a low metastatic capacity led to elevated levels
of p53 protein in clonal transfectants. After intravenous inoculation into syngeneic mice, p53-transfected clones showed significantly
increased metastatic potential in comparison with control transfectants. The observed change did not seem to be due to a change
in growth potential per se since the cell lines showed similar growth properties in vitro.
p53 is a tumor-suppressor protein that can activate and repress transcription. Using the yeast two-hybrid system, we identified two previously uncharacterized human proteins, designated 53BP1 and 53BP2, that bind to p53. 53BP1 shows no significant homology to proteins in available databases, whereas 53BP2 contains two adjacent ankyrin repeats and a Src homology 3 domain. In vitro binding analyses indicate that both of these proteins bind to the central domain of p53 (residues 80-320) required for site-specific DNA binding. Consistent with this finding, p53 cannot bind simultaneously to 53BP1 or 53BP2 and to a DNA fragment containing a consensus p53 binding site. Unlike other cellular proteins whose binding to p53 has been characterized, both 53BP1 and 53BP2 bind to the wild-type but not to two mutant p53 proteins identified in human tumors, suggesting that binding is dependent on p53 conformation. The characteristics of these interactions argue that 53BP1 and 53BP2 are involved in some aspect of p53-mediated tumor suppression.
In an effort to correlate the biological activity of the p53 protein with its conformation, we analysed 14 p53 mutants representative of the most frequently observed protein alterations in human cancers, at codons 175, 248 and 273 (22% of all mutations thus far reported), all three of which contained a CpG dinucleotide. Strikingly, most of the mutants at codons 248 and 273 did not display any change in their conformation, as probed by monoclonal antibodies PAb240 and PAb1620 or by binding to hsp70 protein. For all 14 mutants tested, we found a strict correlation between the transactivation properties of p53, tested either on RGC sequences or using the WAF-1 promoter, and inhibition of cell proliferation. All these mutants showed nuclear localization. Several mutants, present at a low incidence in human tumours, displayed wild-type activity in all our assays, suggesting that the presence of a mutation is not strictly correlated with p53 protein inactivation in tumours. Further analysis of nine thus far undescribed p53 mutants at codon 175 revealed a wild-type or mutant behaviour. All these results suggest that the occurrence of a mutation is dependent on two criteria: (i) the mutability of a given codon, such as those containing a CpG dinucleotide; (ii) the resulting amino acids, eventually leading to synthesis of a p53 conferring a growth advantage on the cell.
The p53 tumor-suppressor gene product is frequently inactivated in malignancies by point mutation. Although most tumor-derived p53 mutants show loss of sequence specific transcriptional activation, some mutants have been identified which retain this activity. One such mutant, p53175P, is defective for the suppression of transformation in rodent cells, despite retaining the ability to suppress the growth of p53-null human cells. We now demonstrate that p53175P can induce a cell-cycle arrest in appropriate cell types but shows loss of apoptotic function. Our results therefore support a direct role of p53 transcriptional activation in mediating a cell-cycle arrest and demonstrate that such activity is not sufficient for the full apoptotic response. These data suggest that either p53 can induce apoptosis through a transcriptionally independent mechanism, a function lost by p53175P, or that this mutant has specifically lost the ability to activate genes which contribute to cell death, despite activation of genes responsible for the G1 arrest. This dissociation of the cell-cycle arrest and apoptotic activities of p53 indicates that inactivation of p53 apoptotic function without concomitant loss of growth inhibition can suffice to relieve p53-dependent tumor-suppression in vivo and thereby contribute to tumor development.
The mechanisms causing resistance to chemotherapeutic drugs in cancer patients are poorly understood. Recent evidence suggests that different forms of chemotherapy may exert their cytotoxic effects by inducing apoptosis. The tumor suppressor gene P53 has a pivotal role inducing apoptosis in response to cellular damage. In vitro investigations have shown intact p53 to play a critical role executing cell death in response to treatment with cytotoxic drugs like 5-fluorouracil, etoposide and doxorubicin. Recently, mutations in the P53 gene were found to confer resistance to anthracyclines in a mouse sarcoma tumor model, and overexpression of the p53 protein (which, in most cases, is due to a mutated gene) was found to be associated with lack of response to cisplatin-based chemotherapy in non-small cell lung cancer. Previous studies have shown mutations in the P53 gene or overexpression of the p53 protein to predict a poor prognosis, but also a beneficial effect of adjuvant radiotherapy or chemotherapy in breast cancer. In this study we present data linking specific mutations in the P53 gene to primary resistance to doxorubicin therapy and early relapse in breast cancer patients.
Human wild-type (wt) p53 can induce apoptosis in transiently transfected H1299 cells maintained at 37 degrees C, whereas tumor-derived
mutant forms of p53 (with the mutation Ala-143, His-175, or Trp-248) fail to do so. At 37 degrees C, p53 with a mutation to
Ala at amino acid 143 (p53Ala143) was transcriptionally inactive. However, at 32 degrees C, p53Ala143 strongly activated transcription
from several physiologically relevant p53-responsive promoters, to extents similar or greater than that of wt p53. Unexpectedly,
p53Ala143 was defective in inducing apoptosis in H1299 cells at 32 degrees C. Concomitantly with the loss of apoptotic activity,
p53Ala143 was found to be deficient in its ability to activate transcription from the p53-responsive portions of the Bax and
insulin-like growth factor-binding protein 3 gene promoters. It is proposed that there may exist distinct classes of p53-responsive
promoters, whose ability to be activated by p53 can be regulated differentially. Such differential regulation may have functional
consequences for the effects of p53 on cell fate.
The involvement of p53 in regulating diverse cellular processes dictates that it must respond to multiple signaling mechanisms, thus coordinating the response to various "stress conditions." Genotoxic stress has served as a paradigm to dissect the transactivation-dependent branch of the pathway by which p53 can induce growth arrest. Alternate mechanisms have been invoked to explain transactivation-independent effects of p53, especially in the context of apoptosis. We have identified a p53-dependent pathway initiated by the gas1 product, a plasma membrane protein highly expressed during G0, which activates a transactivation-independent p53 growth arrest function. Through a detailed deletional analysis and site-specific mutagenesis of p53 we show that the Gas1-dependent signal transduction relies on a proline-rich region (amino acids 63-85) of murine p53. In vivo competition experiments using combinations of such mutants implicate this functional domain of p53 as a docking site in the transmission of antiproliferative signals.
We demonstrate here that synthetic 22-mer peptide 46, corresponding to the carboxy-terminal amino acid residues 361-382 of p53, can activate specific DNA binding of wild-type p53 in vitro and can restore the transcriptional transactivating function of at least some mutant p53 proteins in living cells. Introduction of peptide 46 in Saos-2 cells carrying a Tet-regulatable His-273 mutant p53 construct caused growth inhibition and apoptosis in the presence of mutant p53 but not in its absence, confirming that the effect of the peptide is mediated by reactivation of mutant p53. Moreover, peptide 46 caused apoptosis in mutant as well as wild-type p53-carrying human tumor cell lines of different origin, whereas p53 null tumor cells were not affected. These findings raise possibilities for developing drugs that restore the tumor suppressor function of mutant p53 proteins, thus selectively eliminating tumor cells.
The tumour-suppressor gene TP53 is frequently mutated in breast tumours, and the majority of the mutations are clustered within the core domain, the region involved in DNA binding. We searched for alterations in this central domain of the TP53gene in 222 human breast cancer specimens using polymerase chain reaction-single-strand conformation analysis (PCR-SSCA) followed by sequencing. TP53 gene mutations were observed in 66 tumours (31%), including three tumours that contain two mutations. Fifty-four (78%) of these mutations were missense point mutations, one was a nonsense mutation and four were deletions and/or insertions causing disruption of the protein reading frame, whereas four mutations were either silent or a polymorphism (at codon 213; n = 6). Interestingly, the majority of missense mutations were observed at codon 248. The outcome has been related with patient and tumour characteristics, and with prognosis in 177 patients who were eligible for analysis of both relapse-free and overall survival (median survival for patients alive was 115 months). There was no significant association between the frequency of TP53 mutations and menopausal or nodal status, or tumour size. In a Cox univariate analysis, TP53 gene mutation was significantly associated with poor relapse-free survival (RFS: P = 0.02) but not with overall survival (OS: P = 0.07). In a Cox multivariate analysis, including classical prognostic factors, TP53 gene mutation independently predicted poor RFS and OS (RHR = 1.8 and 1.6 respectively). Unexpectedly, the median relapse-free survival of patients with a polymorphism at codon 213 or with a silent mutation was shorter (median 11 months) than the median relapse-free survival of patients with or without a TP53 gene mutation (median 34 or 48 months respectively). In an exploratory subset analysis, mutations in codons that directly contact DNA were related with the poorest relapse-free (P < 0.05) and overall survival (P < 0.02). These data imply that in the analysis of the prognostic value of TP53, the type of mutation and its biological function should be considered.
Wild-type p53 is a tumor suppressor gene which can activate or repress transcription, as well as induce apoptosis. The human p53 proline-rich domain localized between amino acids 64 and 92 has been reported to be necessary for efficient growth suppression. This study shows that this property mainly results from impaired apoptotic activity. Although deletion of the proline-rich domain does not affect transactivation of several promoters, such as WAF1, MDM2 and BAX, it does alter transcriptional repression, reactive oxygen species production and sequence-specific transactivation of the PIG3 gene, and these are activities which affect apoptosis. Whereas gel retardation assays revealed that this domain did not alter in vitro the specific binding to the p53-responsive element of PIG3, this domain plays a critical role in transactivation from a synthetic promoter containing this element. To explain this discrepancy, evidence is given for a proline-rich domain-mediated cellular activation of p53 DNA binding.
Mutant, but not wild-type p53 binds with high affinity to a variety of MAR-DNA elements (MARs), suggesting that MAR-binding of mutant p53 relates to the dominant-oncogenic activities proposed for mutant p53. MARs recognized by mutant p53 share AT richness and contain variations of an AATATATTT "DNA-unwinding motif," which enhances the structural dynamics of chromatin and promotes regional DNA base-unpairing. Mutant p53 specifically interacted with MAR-derived oligonucleotides carrying such unwinding motifs, catalyzing DNA strand separation when this motif was located within a structurally labile sequence environment. Addition of GC-clamps to the respective MAR-oligonucleotides or introducing mutations into the unwinding motif strongly reduced DNA strand separation, but supported the formation of tight complexes between mutant p53 and such oligonucleotides. We conclude that the specific interaction of mutant p53 with regions of MAR-DNA with a high potential for base-unpairing provides the basis for the high-affinity binding of mutant p53 to MAR-DNA.
The p53 tumor suppressor protein, found mutated in over 50% of all human tumors, is a sequence-specific transcriptional activator. Recent studies have identified a p53 relative, termed p73. We were interested in determining the relative abilities of wild-type and mutant forms of p53 and p73α and -β isoforms to transactivate various p53-responsive promoters. We show that both p73α and p73β activate the transcription of reporters containing a number of p53-responsive promoters in the p53-null cell line H1299. However, a number of significant differences were observed between p53 and p73 and even between p73α and p73β. Additionally, a
Saccharomyces cerevisiae
-based reporter assay revealed a broad array of transcriptional transactivation abilities by both p73 isoforms at 37°C. Recent data have shown that p73 can associate with p53 by the yeast two-hybrid assay. When we examined complex formation in transfected mammalian cells, we found that p73α coprecipitates with mutant but not wild-type p53. Since many tumor-derived p53 mutants are capable of inhibiting transactivation by wild-type p53, we tested the effects of two representative hot-spot mutants (R175H and R248W) on p73. By cotransfecting p73α along with either p53 mutant and a p53-responsive reporter, we found that both R175H and R248W reduces the transcriptional activity of p73α. This decrease in transcriptional activity is correlated with the reduced ability of p73α to promote apoptosis in the presence of tumor-derived p53 mutants. Our data suggest the possibility that in some tumor cells, an outcome of the expression of mutant p53 protein may be to interfere with the endogenous p73 protein.
Many tumors overexpress mutant forms of p53. A growing number of studies suggest that the nature of a p53 mutation in a cell can impact upon cellular properties, clinical responses to therapy and prognosis of a tumor. To explore the cellular basis of these observations, experiments were designed to compare the properties of cells with and without p53 mutations within the same cell population. To that end, various tumor-derived human p53 mutants were introduced into p53-null H1299 lung adenocarcinoma cells. Clonogenic survival assays revealed that cells overexpressing the p53His175 mutant, but not the p53His273 mutant, recover preferentially from etoposide treatment. Moreover, p53His175 as well as p53His179 reduced substantially the rate of etoposide-induced apoptosis, whereas p53His273 and p53Trp248 had a much milder protective effect. In contrast, p53His175 and p53His273 exerted very similar effects on the cellular response to cisplatin; both conferred increased resistance to low concentrations of the drug (2.5 microg/ml), but did not protect at all against high concentrations (10 microg/ml). Hence particular p53 mutants may confer upon tumor cells a selective survival advantage during chemotherapy. These findings define a new type of mutant p53 selective gain of function, which may compromise the efficacy of cancer chemotherapy.
The p53 tumor‐suppressor gene product is frequently inactivated in malignancies by point mutation. Although most tumor‐derived p53 mutants show loss of sequence specific transcriptional activation, some mutants have been identified which retain this activity. One such mutant, p53175P, is defective for the suppression of transformation in rodent cells, despite retaining the ability to suppress the growth of p53‐null human cells. We now demonstrate that p53175P can induce a cell‐cycle arrest in appropriate cell types but shows loss of apoptotic function. Our results therefore support a direct role of p53 transcriptional activation in mediating a cell‐cycle arrest and demonstrate that such activity is not sufficient for the full apoptotic response. These data suggest that either p53 can induce apoptosis through a transcriptionally independent mechanism, a function lost by p53175P, or that this mutant has specifically lost the ability to activate genes which contribute to cell death, despite activation of genes responsible for the G1 arrest. This dissociation of the cell‐cycle arrest and apoptotic activities of p53 indicates that inactivation of p53 apoptotic function without concomitant loss of growth inhibition can suffice to relieve p53‐dependent tumor‐suppression in vivo and thereby contribute to tumor development.
Point mutations in the p53 gene are the most frequently identified genetic change in human cancer. They convert murine p53 from a tumour suppressor gene into a dominant transforming oncogene able to immortalize primary cells and bring about full transformation in combination with an activated ras gene. In both the human and murine systems the mutations lie in regions of p53 conserved from man to Xenopus. We have developed a monoclonal antibody to p53 designated PAb240 which does not immunoprecipitate wild type p53. A series of different p53 mutants all react more strongly with PAb240 than with PAb246. The PAb240 reactive form of p53 cannot bind to SV40 large T antigen but does bind to HSP70. In contrast, the PAb246 form binds to T antigen but not to HSP70. PAb240 recognizes all forms of p53 when they are denatured. It reacts with all mammalian p53 and chicken p53 in immunoblots. We propose that immunoprecipitation of p53 by PAb240 is diagnostic of mutation in both murine and human systems and suggest that the different point mutations which convert p53 from a recessive to a dominant oncogene exert a common conformational effect on the protein. This conformational change abolishes T antigen binding and promotes self-oligomerization. These results are consistent with a dominant negative model where mutant p53 protein binds to and neutralizes the activity of p53 in the wild type conformation.
The aim of our study was to evaluate if p53 mutations, especially those in the L2/L3 domains of the p53 gene, add prognostic information for node-positive and steroid receptor positive breast cancer patients. Two hundred and five tumour samples from a randomised clinical trial of 596 lymph node- and steroid receptor positive breast cancer patients were included. All patients had been randomly allocated to receive 20mg of adjuvant tamoxifen (TAM) daily for 2 years or TAM plus one cycle of low-dose, short-term chemotherapy. For detection of p53 mutations we used in vitro amplification by polymerase chain reaction and consecutively performed temperature gradient gel electrophoresis (PCR-TGGE) and direct sequencing. We found p53 mutations in 42/205 (20%) cases: 16/42 (38%) p53 mutations occurred within the L2/L3 domains of the p53 gene, and 26/42 (62%) outside the L2/L3 domains. p53 mutation served as a statistically significant parameter in predicting disease-free survival in univariate (P=0.02) and multivariate (P=0.009) analysis. For overall survival, no significant differences were observed. Patients with tumours that had p53 mutations within the L2/L3 domains of the gene showed no significant difference to those with mutations outside the L2/L3 domains for disease-free survival. For overall survival, mutations in the L2/L3 domains showed a marginally significant difference (P=0.05) in multivariate analysis, but not in univariate analysis (P=0.13). We conclude that mutation in the L2/L3 domains of the p53 gene is not an independent prognostic indicator of disease outcome for patients suffering from breast cancer with lymph node metastases and positive steroid receptors.
Colorectal cancer can arise as a result of spontaneous genetic lesions or an inherited defect. In this issue, [ Howe et al ][1]. report that the gene defective in one of the inherited syndromes is SMAD4 , a member of a key signal transduction pathway. In their commentary, Kinzler and Vogelstein add this gene to a classification scheme for colorectal cancers that groups the offending genes as "gatekeepers," "caretakers," or "landscapers." Landscaping genes such as SMAD4 have an indirect effect on the tissue that will eventually become cancerous and create an abnormal microenvironment for the cells, probably by acting in the adjacent stromal cells.
[1]: http://www.sciencemag.org/cgi/content/short/280/5366/1086
p53 is the most frequently inactivated tumor suppressor gene in human cancer, whereas its homologue, p73, is rarely mutated.
Similarly to p53, p73 can promote growth arrest or apoptosis when overexpressed in certain p53-null tumor cells. It has previously
been shown that some human tumor-derived p53 mutants can exert gain of function activity. The molecular mechanism underlying
this activity remains to be elucidated. We show here that human tumor-derived p53 mutants (p53His175 and p53Gly281) associate
in vitro andin vivo with p73α, β, γ, and δ. This association occurs under physiological conditions, as verified in T47D and SKBR3 breast cancer
cell lines. The core domain of mutant p53 is sufficient for the association with p73, whereas both the specific DNA binding
and the oligomerization domains of p73 are required for the association with mutant p53. Furthermore, p53His175 and p53Gly281
mutants markedly reduce the transcriptional activity of the various isoforms of p73. Thus, human tumor-derived p53 mutants
can associate with p73 not only physically but also functionally. These findings define a network involving mutant p53 and
the various spliced isoforms of p73 that may confer upon tumor cells a selective survival advantage.
Recent experiments have suggested that p53 action may be mediated through its interaction with DNA. We have now identified 18 human genomic clones that bind to p53 in vitro. Precise mapping of the binding sequences within these clones revealed a consensus binding site with a striking internal symmetry, consisting of two copies of the 10 base pair motif 5'-PuPuPuC(A/T)(T/A)GPyPyPy-3' separated by 0-13 base pairs. One copy of the motif was insufficient for binding, and subtle alterations of the motif, even when present in multiple copies, resulted in loss of affinity for p53. Mutants of p53, representing each of the four "hot spots" frequently altered in human cancers, failed to bind to the consensus dimer. These results define the DNA sequence elements with which p53 interacts in vitro and which may be important for p53 action in vivo.
Wild-type p53 has recently been shown to repress transcription from several cellular and viral promoters. Since p53 mutations are the most frequently reported genetic defects in human cancers, it becomes important to study the effects of mutations of p53 on promoter functions. We, therefore, have studied the effects of wild-type and mutant human p53 on the human proliferating-cell nuclear antigen (PCNA) promoter and on several viral promoters, including the herpes simplex virus type 1 UL9 promoter, the human cytomegalovirus major immediate-early promoter-enhancer, and the long terminal repeat promoters of Rous sarcoma virus and human T-cell lymphotropic virus type I. HeLa cells were cotransfected with a wild-type or mutant p53 expression vector and a plasmid containing a chloramphenicol acetyltransferase reporter gene under viral (or cellular) promoter control. As expected, expression of the wild-type p53 inhibited promoter function. Expression of a p53 with a mutation at any one of the four amino acid positions 175, 248, 273, or 281, however, correlated with a significant increase of the PCNA promoter activity (2- to 11-fold). The viral promoters were also activated, although to a somewhat lesser extent. We also showed that activation by a mutant p53 requires a minimal promoter containing a lone TATA box. A more significant increase (25-fold) in activation occurs when the promoter contains a binding site for the activating transcription factor or cyclic AMP response element-binding protein. Using Saos-2 cells that do not express p53, we showed that activation by a mutant p53 was a direct enhancement. The mutant forms of p53 used in this study are found in various cancer cells. The activation of PCNA by mutant p53s may indicate a way to increase cell proliferation by the mutant p53s. Thus, our data indicate a possible functional role for the mutants of p53 found in cancer cells in activating several important loci, including PCNA.
Point mutations in the p53 gene are the most frequently identified genetic change in human cancer. They convert murine p53 from a tumour suppressor gene into a dominant transforming oncogene able to immortalize primary cells and bring about full transformation in combination with an activated ras gene. In both the human and murine systems the mutations lie in regions of p53 conserved from man to Xenopus. We have developed a monoclonal antibody to p53 designated PAb240 which does not immunoprecipitate wild type p53. A series of different p53 mutants all react more strongly with PAb240 than with PAb246. The PAb240 reactive form of p53 cannot bind to SV40 large T antigen but does bind to HSP70. In contrast, the PAb246 form binds to T antigen but not to HSP70. PAb240 recognizes all forms of p53 when they are denatured. It reacts with all mammalian p53 and chicken p53 in immunoblots. We propose that immunoprecipitation of p53 by PAb240 is diagnostic of mutation in both murine and human systems and suggest that the different point mutations which convert p53 from a recessive to a dominant oncogene exert a common conformational effect on the protein. This conformational change abolishes T antigen binding and promotes self-oligomerization. These results are consistent with a dominant negative model where mutant p53 protein binds to and neutralizes the activity of p53 in the wild type conformation.
Although the case for p53 as a tumor suppressor gene appears very strong, one should still keep an open eye for the possibility that mutations in p53 do not necessarily imply a mere loss of "suppressor" activity. It is still possible that the presence of a p53 mutation in a tumor contributes, in a dominant positive manner, to tumorigenesis. In other words, certain p53 mutants may well be oncogenic in their own right, and carry distinct activities that promote growth deregulation and malignant progression. Elucidating this issue also has practical implications, since the nature of the resident mutations may greatly dictate the consequences of attempts to reintroduce wild-type (wt) p53 into particular types of tumor cells. There are two major obstacles along the road to meaningful answers: the limitations of the experimental systems used for evaluating the biological activities of wt and mutant p53 and a fundamental lack of knowledge about the relevant biochemistry of the p53 protein. These two aspects constitute primary experimental challenges for investigators in the field.
In its wild-type form, the protein p53 can interfere with neoplastic processes. Tumor-derived cells often express mutant p53. Full-length mutant forms of p53 isolated so far from transformed mouse cells exhibit three common properties in vitro: loss of transformation-suppressing activity, gain of pronounced transforming potential, and ability to bind the heat shock protein cognate hsc70. A tumor-derived mouse p53 variant is now described, whose site of mutation corresponds to a hot spot for p53 in human tumors. While absolutely nonsuppressing, it is only weakly transforming and exhibits no detectable hsc70 binding. The data suggest that the ability of a p53 mutant to bind endogenous p53 is not the sole determinant of its oncogenic potential. The data also support the existence of gain-of-function p53 mutants.
The majority of the p53 genes derived from human colorectal carcinomas contain point mutations. A significant number of these mutations occur in or around amino acids 143, 175, 273, or 281. Experiments presented here demonstrate for the first time that p53 DNA clones containing any one of these mutations cooperate with the activated ras oncogene to transform primary rat embryo cells in culture. These transformed cells produce elevated levels of the human p53 protein, which has extended half-lives (1.5-7 h), as compared to the wild-type human p53 protein (20-30 min). The p53 mutant with an alteration at residue 175 (p53-175H) binds tightly to the cellular heat shock protein, hsc70. In contrast, the p53 mutants possessing mutations at either residue 273 or 281 (p53-273H/281G) do not bind detectably to this heat shock protein and generally are less efficient at forming transformed foci in culture. The transformed cell lines are tumorigenic in nude mice. Thus, two classes of p53 mutant proteins can be distinguished: p53-175H, which cooperates with ras efficiently and binds to hsc70, and p53-273H/281G, which has a reduced efficiency of transformed foci formation and does not bind hsc70. This demonstrates that complex formation between mutant p53 and hsc70 is not required for p53-mediated transformation, but rather it facilitates this function, perhaps by ensuring sequestration of the endogenous wild-type p53 protein. The positive effect on cell proliferation by these mutant p53 proteins is consistent with a role for activated p53 mutants in the genesis of colorectal carcinomas.
Human p53 displays two immunodominant regions localized in the amino and carboxy termini of the protein. Using a truncated p53 (residues 66 to 361), we selected eight new monoclonal antibodies directed to the central part of the protein. We identified the epitopes recognized by seven out of eight antibodies with a set of overlapping peptides. One of these antibodies had an epitope similar to PAb240, whereas the others recognized novel and diverse antigenic determinants. Using a series of 19 p53 mutants, we show that the behavior of several of the new monoclonal antibodies is similar to that of PAb240 despite their various epitope localizations. This suggests that different mutations in the p53 protein induce an overall conformational change that can be detected by various monoclonal antibodies directed toward the central part of the protein.
We have examined the relationship between point mutation of the p53 tumor suppressor gene and survival in colorectal cancer patients. We found that patients with tumors harboring mutated p53 genes showed a significantly poorer prognosis than did those patients with genes without point mutations, and, moreover, patient response to postoperative therapies depended significantly on mutation status in both adjuvant and palliative treatment cohorts. However, not all point mutations were the same functionally; point mutations within the conserved domains of the p53 tumor suppressor gene were inherently more aggressive than tumors with point mutations outside of these domains, and mutations of codon 175 were particularly aggressive. These results suggest that knowledge of a patient's p53 status, both with respect to the presence of point mutations and to the specific nature of the lesion, may be required to accurately predict both the course of the disease and the response of the disease to postoperative therapeutic interventions, especially those therapies based on the induction of apoptosis in the neoplastic cell.
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The p53 tumor suppressor gene is mutated in a large percentage of human malignancies, including tumors of the colon, breast, lung and brain. Individuals who inherit one mutant allele of p53 are susceptible to a wide range of tumor types. The gene encodes a transcriptional regulator that may function in the cellular response to DNA damage. The construction of mouse strains carrying germline mutations of p53 facilitates analysis of the function of p53 in normal cells and tumorigenesis.
In order to study the effects of p53 mutation in vivo, we have constructed a mouse strain carrying a germline disruption of the gene. This mutation removes approximately 40% of the coding capacity of p53 and completely eliminates synthesis of p53 protein. As observed previously for a different germline mutation of p53, animals homozygous for this p53 deletion mutation are viable but highly predisposed to malignancy. Heterozygous animals also have an increased cancer risk, although the distribution of tumor types in these animals differs from that in homozygous mutants. In most cases, tumorigenesis in heterozygous animals is accompanied by loss of the wild-type p53 allele.
We reaffirm that p53 function is not required for normal mouse development and conclude that p53 status can strongly influence tumor latency and tissue distribution.
Mutations in the p53 tumor suppressor are the most frequently observed genetic alterations in human cancer. The majority of
the mutations occur in the core domain which contains the sequence-specific DNA binding activity of the p53 protein (residues
102-292), and they result in loss of DNA binding. The crystal structure of a complex containing the core domain of human p53
and a DNA binding site has been determined at 2.2 angstroms resolution and refined to a crystallographic R factor of 20.5
percent. The core domain structure consists of a beta sandwich that serves as a scaffold for two large loops and a loop-sheet-helix
motif. The two loops, which are held together in part by a tetrahedrally coordinated zinc atom, and the loop-sheet-helix motif
form the DNA binding surface of p53. Residues from the loop-sheet-helix motif interact in the major groove of the DNA, while
an arginine from one of the two large loops interacts in the minor groove. The loops and the loop-sheet-helix motif consist
of the conserved regions of the core domain and contain the majority of the p53 mutations identified in tumors. The structure
supports the hypothesis that DNA binding is critical for the biological activity of p53, and provides a framework for understanding
how mutations inactivate it.
We report that the expression of murine or human mutant p53 proteins in cells with no endogenous p53 proteins confers new or additional phenotypes upon these cells. Mutant p53 proteins expressed in cell lines lacking p53 resulted in either enhanced tumorigenic potential in nude mice ((10)3 cells) or enhanced plating efficiency in agar cell culture (human SAOS-2 cells). Also, mutant human p53 alleles, unlike the wild-type p53 protein, could also enhance the expression of a test gene regulated by the multi-drug resistance enhancer-promoter element. These data demonstrate a gain of function associated with p53 mutations in addition to the loss of function shown previously to be associated with mutations in this tumour suppressor gene.
Acquired mutations in TP53 as well as immunohistochemically detectable protein expression have been implicated as prognostic factors for breast cancer. We have evaluated the relationship between mutations detected in 119 breast tumours and various clinicohistopathological indices, stratifying the mutations according to the functional domains as defined by the recent elucidation of the crystal structure of the protein. Patients with missense mutations located in regions encoding parts of the protein involved in zinc-binding had significantly decreased disease-free and overall survival relative to patients whose tumours had mutations in other domains. These results indicate that these biochemically defined domains also have biological relevance in terms of breast cancer disease course, and suggest that some mutations in TP53, more than others, can contribute to the development of clinically more aggressive and perhaps treatment resistant breast tumours. When confirmed, this will be of potential importance in predicting the clinical behaviour of breast cancer and its responsiveness to therapy.
The gene encoding the tumour suppressor protein p53 is one of the most commonly mutated genes in human cancers. Analysis of the mutational events that target the p53 gene has revealed evidence for both exogenous and endogenous mutational mechanisms. For example, the p53 mutational spectrum reveals evidence for a direct causal effect of ultraviolet radiation in skin cancer, of aflatoxin B1 in liver cancer and of tobacco smoke in lung cancer. This novel field, molecular epidemiology of human cancer risk, has added a new dimension to classical associative epidemiology by providing a direct link between human cancer and carcinogen exposure.
Mutations in the p53 tumor suppressor are among the most frequently observed genetic alterations in human cancer and map to the 200-amino acid core domain of the protein. The core domain contains the sequence-specific DNA binding activity and the in vitro 53BP2 protein binding activity of p53. The crystal structure of the p53 core domain bound to the 53BP2 protein, which contains an SH3 (Src homology 3) domain and four ankyrin repeats, revealed that (i) the SH3 domain binds the L3 loop of p53 in a manner distinct from that of previously characterized SH3-polyproline peptide complexes, and (ii) an ankyrin repeat, which forms an L-shaped structure consisting of a beta hairpin and two alpha helices, binds the L2 loop of p53. The structure of the complex shows that the 53BP2 binding site on the p53 core domain consists of evolutionarily conserved regions that are frequently mutated in cancer and that it overlaps the site of DNA binding. The six most frequently observed p53 mutations disrupt 53BP2 binding in vitro. The structure provides evidence that the 53BP2-p53 complex forms in vivo and may have a critical role in the p53 pathway of tumor suppression.
Activation of the p53 tumor suppressor protein has been demonstrated to block cell growth by inducing either a transient cell cycle arrest or programmed cell death (apoptosis). Although evidence exists linking p53's function as an activator of transcription to its ability to effect cell cycle arrest, the role of this activity in the induction of apoptosis remains unclear. To gain insight into the molecular mechanisms underlying p53-mediated antiproliferative pathways, a study was initiated to explore the functions of a putative p53 signaling domain. This region of the human p53 protein is localized between amino acids 61 and 94 (out of 393) and is noteworthy in that it contains five repeats of the sequence PXXP (where P represents proline and X any amino acid). This motif has been shown to play a role in signal transduction via its SH3 domain binding activity. A p53 cDNA deletion mutant (delta pro AE), which lacks this entire proline-rich domain (deleted for amino acids 62-91), was created and characterized for a variety of p53 functions. The entire domain has been shown to be completely dispensable for transcriptional activation. On the other hand, this deletion of the p53 proline-rich domain impairs p53's ability to suppress tumor cell growth in culture. Amino acid substitution mutations at residues 22 and 23 of p53 (eliminates transcriptional activity) also impair p53-mediated inhibition of cell growth in culture. Unlike wild-type p53, the delta proAE mutant cDNA can be stably expressed in tumor derived cell lines with few immediate detrimental effects. These cells express physiologic levels of p53 protein that are induced normally in response to DNA damage, indicating that removal of the proline-rich domain does not disrupt p53's upstream regulation by DNA damage. These data indicate that, in addition to the transcriptional activation domain, the p53 proline-rich domain plays a critical role in the transmission of antiproliferative signals down-stream of the p53 protein and may link p53 to a direct signal transduction pathway.
In a consecutive series of 222 colorectal carcinomas from patients with a median follow-up time of 56.8 months (range, 0.5-92.2) treated with surgery, the TP53 gene was screened for mutations. Exons 5-8 were analyzed using constant denaturant gel electrophoresis followed by sequencing, and mutations were found in 102 cases (45.9%). Mutations were found more frequently in rectal tumors versus other locations (P = 0.029) and in aneuploid compared to diploid tumors (P < 0.001). Presence of a TP53 mutation was also significantly associated with absence of microsatellite instability (P = 0.028), as well as with loss of heterozygosity at 17p13 (P < 0.001). The TP53 mutations in the left-sided and rectal tumors were more often transversions than transitions, indicating a different etiology in the development of these tumors. The tendency for shorter cancer-related survival among patients with mutations in their tumors was only statistically significant for patients with left-sided tumors (P = 0.003). All patients with mutations affecting the L3 domain of the protein involved in zinc binding had a significantly shorter cancer-related survival (P = 0.036), indicating that mutations affecting this domain have biological relevance in terms of colorectal cancer disease course. These results suggest that knowledge of a patient's TP53 status, with respect to both the presence and the localization of the mutation, may be important in prognosis evaluation, particularly when selecting patients for more aggressive postoperative therapeutic intervention.
Tumor suppressors act along diverse biochemical pathways to function as safeguards against cancer. This review summarizes how these pathways can be regulated, primarily by focusing on the well-characterized wild-type p53 tumor suppressor as a paradigm. Specifically, we discuss recent data linking p53 to the processes of signal transduction and angiogenesis.