p63 heterozygous mutant mice are not prone to spontaneous or chemically induced tumors

Homology between p63 and p53 has suggested that these proteins might function similarly. However, the majority of data from human tumors have not supported a similar role for p63 in tumor suppression. To investigate this issue, we studied spontaneous tumorigenesis in p63+/- mice in both WT and p53-compromised backgrounds. We found that p63+/- mice were not tumor prone and mice heterozygous for both p63 and p53 had fewer tumors than p53+/- mice. The rare tumors that developed in mice with compromised p63 were also distinct from those of p53+/- mice. Furthermore, p63+/- mice were not prone to chemically induced tumorigenesis, and p63 expression was maintained in carcinomas. These findings demonstrate that, in agreement with data from human tumors, p63 plays a markedly different biological role in cancer than p53.     

Multiple primary tumors as an indicator for p16INK4a germline mutations in pancreatic cancer patients?

Multiple primary tumors in pancreatic cancer patients might indicate a genetic predisposition to the development of malignancies. In this study we evaluated whether the mutation rate of the TP53 and p16INK4a genes of pancreatic cancers differs in pancreatic cancer patients with and without multiple primaries. Furthermore, we investigated whether pancreatic cancer patients with multiple primaries carry germline mutations in either p16INK4a, TP53, or BRCA2 tumor suppressor genes to detect a genetic alteration that predisposes to the development of different primaries. Fourteen (23%) of 60 pancreatic cancer patients developed histologically verified additional primaries during their lifetimes. Normal constitutional and tumor DNA of the 14 patients with a positive cancer history, but negative family history, were analyzed for p16INK4a, TP53, and BRCA2 mutations by single-strand conformational variant (SSCV) analysis and direct sequencing. Hypermethylation of the p16INK4a promoter region in pancreatic cancers was identified by methylation-specific polymerase chain reaction (PCR; MSP). Four of 14 pancreatic carcinomas carried somatic intragenic p16INK4a mutations, and another four tumors revealed hypermethylation of the p16INK4a promoter region. Somatic intragenic TP53 mutations were identified in six of 14 tumors. None of the pancreatic cancer patients carried TP53 or BRCA2 germline mutations. In contrast, one of 14 pancreatic cancer patients with multiple primaries carried the p16INK4a mutation A68V in his germline. This mutation was localized in the conserved second ankyrin repeat of p16INK4a and did not occur in 100 control patients. The frequency of somatic TP53 and p16INK4a mutations in pancreatic cancer is similar in patients with and without multiple primaries. TP53 and BRCA2 germline mutations seem not to be significantly associated with the occurrence of multiple primaries in pancreatic cancer patients. However, p16INK4a germline mutations might be causative for tumor development in some pancreatic cancer patients with multiple primaries. The genetic investigation of patients with accumulation of different cancers even without a positive family history may be a new approach for the understanding of the relation of different cancers.     

Methylation status analysis of cell cycle regulatory genes (p16INK4A, p15INK4B, p21Waf1/Cip1, p27Kip1 and p73) in natural killer cell disorders

Natural killer (NK) cell disorders are rare diseases. Genetic abnormalities of the several tumor suppressor genes, including p15INK4B, p16INK4A/p14ARF, p53, p73, and Rb genes have been reported. Deletions and point mutations of these genes are frequently detected in these diseases. It has been reported that tumor suppressor genes are inactivated by DNA methylation of the promoter region and/or first exon of the genes in a variety of human cancers. In this study we analyze the methylation status of the genes associated with cell cycle regulation, including p16INK4A, p15INK4B, p21/Waf1/Cip1, p27/Kip1, p73, and p14ARF, by methylation specific (MS) PCR and/or bisulfite sequencing. We examined 29 cases of NK cell disorders (five aggressive NK cell leukemia/lymphoma, three blastic NK cell lymphoma/leukemia, five nasal NK cell lymphoma, three myeloid/NK cell precursor acute leukemia, 13 chronic NK lymphocytosis). We found methylation of the first exon of the p16INK4A gene in two cases (one aggressive, one blastic), and methylation of the p14ARF gene in one aggressive NK cell leukemia. Bisulfite sequencing revealed that methylation of the p15 and p27 genes was rare in these disorders. MS-PCR suggested that the p73 and p21 genes were methylated in seven cases, respectively (p73: one blastic, one nasal, five chronic; p21: one myeloid/NK, one aggressive, one nasal, and four chronic); bisulfite sequencing confirmed that methylated alleles of these genes were dominant in the samples except three cases (one myeloid/NK, one aggressive, and one chronic) in which methylated alleles of the p21 genes were less than 34% of all alleles. These results suggested that inactivation of the cell cycle regulatory genes by DNA methylation could be associated with tumorigenesis in NK cell disorders, not only aggressive subtypes but also chronic subtype.     

Use of p53 transgenic mice in the development of cancer models for multiple purposes

The tumor suppressor gene p53 is perhaps the most commonly mutated gene in human cancer, being mutated in a high percentage of colon, breast, skin, bladder, and many cancers of the aerodigestive tract. Individuals with Li-Fraumeni syndrome, who routinely have a germline mutation in the p53 tumor suppressor gene, are at high risk for lung cancer, confirming its intimate role in lung tumorigenesis in humans. In contrast, the majority of chemically induced or spontaneous cancers in rodents do not contain mutations in p53. Therefore, we examined a transgenic mouse that contains a dominant negative mutation (Arg135Val) in the p53 gene placed under the control of its own endogenous promoter. The resulting mice have 3 copies of the mutated transgene as well as 2 normal p53 alleles. In the chemical carcinogenesis studies, we employed mice containing the mutated p53 gene to examine for carcinogen susceptibility. We found that mice with the p53 mutation, on an A/J F1 background, were more susceptible to a number of potential lung carcinogens, including N-methyl-N-nitrosourea (MNU) and the known tobacco carcinogens 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and benzo(a)pyrene (BP). Mice with a mutant p53 developed larger tumors and roughly 3 times as many tumors, emphasizing the potential effects of a p53 mutation both on tumor initiation and progression. In addition, we examined 2 nonlung carcinogens, 1,2-dimethylhydrazine (DMH), a colon carcinogen, and N-butyl-N-(4-hydroxybutyl)-nitrosamine (OHBBN), a bladder carcinogen. Interestingly a germline p53 mutation increased the incidence of DMH-induced colon, lung, hepatic, and uterine tumors, while having limited effects on OHBBN-induced bladder tumors. Because of its heightened susceptibility we are examining the use of this model in smoke-induced tumorigenesis in A/J mice as well. Employing the lung adenomas induced by NNK, we found that mice with or without a p53 mutation were equally susceptible to the chemopreventive effects of dexamethasone plus myo-inisitol and green tea. These tumors, which arise in a highly reproducible manner in p53 transgenic mice following carcinogen treatment, have mutations in both p53 and the K-ras oncogene. Thus, this model appears useful for examining for potential chemotherapeutic agents. p53-mutated or wild-type mice were equally susceptible to the therapeutic effects of Taxol or Adriamycin. Interestingly, piroxicam was similarly effective in inhibiting colon tumor formation by DMH in mice with or without a mutation in the p53 tumor suppressor gene. In contrast, lung and uterine tumors developing in these mice were not susceptible to the chemopreventive effects of piroxicam. In summary, mice with mutations in the p53 tumor suppressor gene appear to be particularly applicable for basic mechanistic studies, for screening for potential carcinogens, and for screening for chemopreventive or chemotherapeutic agents.     

Genome-wide analysis reveals recurrent structural abnormalities of TP63 and other p53-related genes in peripheral T-cell lymphomas

Peripheral T-cell lymphomas (PTCLs) are aggressive malignancies of mature T lymphocytes with 5-year overall survival rates of only ～ 35%. Improvement in outcomes has been stymied by poor understanding of the genetics and molecular pathogenesis of PTCL, with a resulting paucity of molecular targets for therapy. We developed bioinformatic tools to identify chromosomal rearrangements using genome-wide, next-generation sequencing analysis of mate-pair DNA libraries and applied these tools to 16 PTCL patient tissue samples and 6 PTCL cell lines. Thirteen recurrent abnormalities were identified, of which 5 involved p53-related genes (TP53, TP63, CDKN2A, WWOX, and ANKRD11). Among these abnormalities were novel TP63 rearrangements encoding fusion proteins homologous to ΔNp63, a dominant-negative p63 isoform that inhibits the p53 pathway. TP63 rearrangements were seen in 11 (5.8%) of 190 PTCLs and were associated with inferior overall survival; they also were detected in 2 (1.2%) of 164 diffuse large B-cell lymphomas. As TP53 mutations are rare in PTCL compared with other malignancies, our findings suggest that a constellation of alternate genetic abnormalities may contribute to disruption of p53-associated tumor suppressor function in PTCL.     

Understanding the function-structure and function-mutation relationships of p53 tumor suppressor protein by high-resolution missense mutation analysis

Inactivation of the tumor suppressor p53 by missense mutations is the most frequent genetic alteration in human cancers. The common missense mutations in the TP53 gene disrupt the ability of p53 to bind to DNA and consequently to transactivate downstream genes. However, it is still not fully understood how a large number of the remaining mutations affect p53 structure and function. Here, we used a comprehensive site-directed mutagenesis technique and a yeast-based functional assay to construct, express, and evaluate 2,314 p53 mutants representing all possible amino acid substitutions caused by a point mutation throughout the protein (5.9 substitutions per residue), and correlated p53 function with structure- and tumor-derived mutations. This high-resolution mutation analysis allows evaluation of previous predictions and hypotheses through interrelation of function, structure and mutation.     

Expression of the p53 homologue p63alpha and DeltaNp63alpha in the neoplastic sequence of Barrett's oesophagus: correlation with morphology and p53 protein

BACKGROUND: While loss of p53 function is a key oncogenic step in human tumorigenesis, mutations of p53 are generally viewed as late events in the metaplasia-dysplasia-adenocarcinoma sequence of Barrett's oesophagus. Recent reports of a series of genes (p63, p73, and others) exhibiting close homology to p53 raise the possibility that abnormalities of these p53 family members may exert their influence earlier in the sequence. AIM: Following recent characterisation of expression of p63 and a major isoform DeltaNp63 by generation of an antiserum that recognises p63 isoforms, but not p53, our aim was a comparative study of expression of p63 protein and p53 protein in a morphologically well defined biopsy series representative of all stages of the metaplasia-dysplasia-carcinoma sequence in Barrett's oesophagus. METHODS: A series of 60 biopsy cases representing normal oesophagus through to invasive adenocarcinoma were stained, using immunohistochemistry, with antibodies to p63 and p53. All biopsies derived from patients with endoscopic and histopathological substantiation of a diagnosis of traditional/classical Barrett's oesophagus. RESULTS: There was exact concordance in p53 and p63 expression in more advanced forms of neoplasia, high grade dysplasia, and invasive adenocarcinoma, while p63, but not p53, was detected in the proliferative compartment of some non-neoplastic oesophageal tissue, in both squamous mucosa and in the non-neoplastic metaplastic glandular epithelium. CONCLUSIONS: In neoplastic Barrett's oesophagus there is upregulation of both p63 and p53 while p63 isoforms may well have an important role in epithelial biology in both non-metaplastic and metaplastic mucosa of the oesophagus. While abnormalities of p53 function represent an indisputable and critical element of neoplastic transformation, other closely linked genes and their proteins have a role in both the physiology and pathophysiology of the oesophageal mucosa.     

Somatic loss of BRCA1 and p53 in mice induces mammary tumors with features of human BRCA1-mutated basal-like breast cancer

Women carrying germ-line mutations in BRCA1 are strongly predisposed to developing breast cancers with characteristic features also observed in sporadic basal-like breast cancers. They appear as high-grade tumors with high proliferation rates and pushing borders. On the molecular level, they are negative for hormone receptors and ERBB2, display frequent TP53 mutations, and express basal epithelial markers. To study the role of BRCA1 and P53 loss of function in breast cancer development, we generated conditional mouse models with tissue-specific mutation of Brca1 and/or p53 in basal epithelial cells. Somatic loss of both BRCA1 and p53 resulted in the rapid and efficient formation of highly proliferative, poorly differentiated, estrogen receptor-negative mammary carcinomas with pushing borders and increased expression of basal epithelial markers, reminiscent of human basal-like breast cancer. BRCA1- and p53-deficient mouse mammary tumors exhibit dramatic genomic instability, and their molecular signatures resemble those of human BRCA1-mutated breast cancers. Thus, these tumors display important hallmarks of hereditary breast cancers in BRCA1-mutation carriers.     

Loss of p53 synthesis in zebrafish tumors with ribosomal protein gene mutations

Zebrafish carrying heterozygous mutations for 17 different ribosomal protein (rp) genes are prone to developing malignant peripheral nerve sheath tumors (MPNSTs), a tumor type that is seldom seen in laboratory strains of zebrafish. Interestingly, the same rare tumor type arises in zebrafish that are homozygous for a loss-of-function point mutation in the tumor suppressor gene p53. For these reasons, and because p53 is widely known to be mutated in the majority of human cancers, we investigated the status of p53 in the rp^+/− MPNSTs. Using monoclonal antibodies that we raised to zebrafish p53, we found that cells derived from rp^+/− MPNSTs are significantly impaired in their ability to produce p53 protein even in the presence of a proteasome inhibitor and γ-irradiation. Although the coding regions of the p53 gene remain wild type, the gene is transcribed, and overall protein production rates appear normal in rp^+/− MPNST cells, p53 protein does not get synthesized. This defect is observed in all MPNSTs we examined that were derived from our 17 zebrafish lines with rp gene mutations. To date, studies of p53 in malignancies have focused predominantly on either p53 gene mutations or the aberrant posttranslational regulation of the p53 protein. Our results show that the appropriate amount of numerous ribosomal proteins is required for p53 protein production in vivo and that disruption of this regulation most likely contributes to tumorigenesis.     

Frequent mutation of the p53 gene in human esophageal cancer.

Sequence alterations in the p53 gene have been detected in human tumors of the brain, breast, lung, and colon, and it has been proposed that p53 mutations spanning a major portion of the coding region inactivate the tumor suppressor function of this gene. To our knowledge, neither transforming mutations in oncogenes nor mutations in tumor suppressor genes have been reported in human esophageal tumors. We examined four human esophageal carcinoma cell lines and 14 human esophageal squamous cell carcinomas by polymerase chain reaction amplification and direct sequencing for the presence of p53 mutations in exons 5, 6, 7, 8, and 9. Two cell lines and five of the tumor specimens contained a mutated allele (one frameshift and six missense mutations). All missense mutations detected occurred at G.C base pairs in codons at or adjacent to mutations previously reported in other cancers. The identification of aberrant p53 gene alleles in one-third of the tumors we tested suggests that mutations at this locus are common genetic events in the pathogenesis of squamous cell carcinomas of the esophagus.     

Role of AXL in invasion and drug resistance of colon and breast cancer cells and its association with p53 alterations

AIM To characterize AXL receptor tyrosine kinase(AXL)expression in relationship to tumor protein P53(TP53gene,p53 protein)and its role in tumor invasion and response to therapy.METHODS We used 14 cell lines,including 3 isogenic pairs carrying mutant/knockout p53,to gain insight into the relationship between AXL and TP53.These included HCT116,HCT116.p53 mutant,RKO,and RKO.p53-/-lines(all from colon cancers)as well as breast cancer cell lines MCF7 and 1001(MCF7-p53 mutant clone).He La cell line was used as a positive control for epithelial to mesenchymal transition(EMT).AXL expression was determined by Western blotting using rabbit monoclonal antibody clone C89E7.AXL si RNA silencing was performed and followed by collagen invasion assay.Cell viability analysis using the sulforhodamine B assay and the invasion assay were performed after exposure to chemotherapeutic agents(doxorubicin for breast cancer cells;5FU or irinotecan for colon cancer cells).RESULTS We showed that the introduction of p53 mutations or knockout increased expression levels of AXL in isogenic cells compared to the matching p53 wild-type parental cells.Overall,we found a trend for correlation between the potential EMT candidate AXL,p53 alterations,and EMT markers in colorectal and breast cancers.The expression of AXL in RKO cells,a rare colon cancer cell line with inactive Wnt signaling,suggests that the AXL oncogene might provide an alternative genetic pathway for colorectal carcinogenesis in the absence of Wnt signaling activation and TP53 mutation.AXL silencing in the TP53 mutant isogenic cell lines 1001,HCT116.p53 mutant and RKO.P53-/-was95%efficient and the silenced cells were less invasive compared to the parental TP53 wild-type cells.AXL silencing showed a subtle trend to restore colon cancer cell sensitivity to5FU or irinotecan.Importantly,AXL expressing cells developed more invasive potential after exposure to chemotherapy compared to the AXL-silenced cells.CONCLUSION AXL is influenced by p53 status and could cause the emergence of aggressive clones after exposure to chemotherapy.These findings could have applications in cancer management.     

Germ-line mutations of the p53 tumor suppressor gene in patients with high risk for cancer inactivate the p53 protein.

Germ-line mutations in the p53 tumor suppressor gene have been observed in patients with Li-Fraumeni syndrome, brain tumors, second malignancies, and breast cancers. It is unclear whether all of these mutations have inactivated p53 and thereby provide an increased risk for cancer. Therefore, it is necessary to establish the biological significance of these germ-line mutations by the functional and structural analysis of the resulting mutant p53 proteins. We analyzed the ability of seven germ-line mutant proteins observed in patients with Li-Fraumeni syndrome, second primary neoplasms, or familial breast cancer to block the growth of malignant cells and compared the structural properties of the mutant proteins to that of the wild-type protein. Six of seven missense mutations disrupted the growth inhibitory properties and structure of the wild-type protein. One germ-line mutation retained the features of the wild-type p53. Genetic analysis of the breast cancer family in which this mutation was observed indicated that this germ-line mutation was not associated with the development of cancer. These results demonstrate that germ-line p53 mutations observed in patients with Li-Fraumeni syndrome and with second malignancies have inactivated the p53 tumor suppressor gene. The inability of the germ-line p53 mutants to block the growth of malignant cells can explain why patients with these germ-line mutations have an increased risk for cancer. The observation of a functionally silent germ-line mutation indicates that, before associating a germ-line tumor suppressor gene mutation with cancer risk, it is prudent to consider its functional significance.     

Excessive genomic DNA copy number variation in the Li-Fraumeni cancer predisposition syndrome

DNA copy number variations (CNVs) are a significant and ubiquitous source of inherited human genetic variation. However, the importance of CNVs to cancer susceptibility and tumor progression has not yet been explored. Li-Fraumeni syndrome (LFS) is an autosomal dominantly inherited disorder characterized by a strikingly increased risk of early-onset breast cancer, sarcomas, brain tumors and other neoplasms in individuals harboring germline TP53 mutations. Known genetic determinants of LFS do not fully explain the variable clinical phenotype in affected family members. As part of a wider study of CNVs and cancer, we conducted a genome-wide profile of germline CNVs in LFS families. Here, by examining DNA from a large healthy population and an LFS cohort using high-density oligonucleotide arrays, we show that the number of CNVs per genome is well conserved in the healthy population, but strikingly enriched in these cancer-prone individuals. We found a highly significant increase in CNVs among carriers of germline TP53 mutations with a familial cancer history. Furthermore, we identified a remarkable number of genomic regions in which known cancer-related genes coincide with CNVs, in both LFS families and healthy individuals. Germline CNVs may provide a foundation that enables the more dramatic chromosomal changes characteristic of TP53-related tumors to be established. Our results suggest that screening families predisposed to cancer for CNVs may identify individuals with an abnormally high number of these events.     

Ras oncogene and p53 gene hotspot mutations in colorectal cancers

Abstract Ras oncogene and p53 gene mutations are frequently observed in colorectal cancers. The role of co-operation between these two genes in the tumorigenesis of colorectal cancer was evaluated. Point mutations in K-ras oncogene and hotspot codons of p53 gene of colorectal cancers were evaluated by naturally created or amplified created restriction site method. Nine of 42 cases (21.4%) of colorectal cancer showed K-ras oncogene mutations. Six of 42 cases (14.3%) of colorectal cancer showed p53 gene hotspot point mutations. The low frequency of p53 gene mutation in this series may be due to racial difference or different hotspot codons. When six cases with mutated p53 gene were examined, only one (16.7%) showed concurrent K-ras oncogene codon 12 and p53 gene codon 248 mutations. We concluded that the co-operation between ras oncogene and p53 gene hotspot point mutations in the tumorigenesis of colorectal cancer in Chinese was not common. Other factors such as adenomatous polyposis coli gene mutations, oncogene activation or tumour suppression gene inactivation may be involved.     

NDRG1 links p53 with proliferation-mediated centrosome homeostasis and genome stability

The tumor protein 53 (TP53) tumor suppressor gene is the most frequently somatically altered gene in human cancers. Here we show expression of N-Myc down-regulated gene 1 (NDRG1) is induced by p53 during physiologic low proliferative states, and mediates centrosome homeostasis, thus maintaining genome stability. When placed in physiologic low-proliferating conditions, human TP53 null cells fail to increase expression of NDRG1 compared with isogenic wild-type controls and TP53 R248W knockin cells. Overexpression and RNA interference studies demonstrate that NDRG1 regulates centrosome number and amplification. Mechanistically, NDRG1 physically associates with γ-tubulin, a key component of the centrosome, with reduced association in p53 null cells. Strikingly, TP53 homozygous loss was mutually exclusive of NDRG1 overexpression in over 96% of human cancers, supporting the broad applicability of these results. Our study elucidates a mechanism of how TP53 loss leads to abnormal centrosome numbers and genomic instability mediated by NDRG1.Tumor protein 53 (TP53) encodes p53 and is one of the most well-studied tumor suppressor genes. Among its many functions, p53 plays a central role in apoptosis, cell cycle arrest, and maintaining genomic integrity (1–3). Normally, p53 acts as a critical cellular checkpoint monitor in response to stress such as DNA damage. This function prevents cells with aberrant or damaged DNA from proceeding through the cell cycle, allowing time to correct damaged DNA or induce apoptosis if DNA cannot be repaired. This critical role thus prevents cells with altered DNA from inappropriate cell division.Although TP53 is one of the most well described tumor suppressor genes, the mechanisms of many of its functions have not been fully elucidated. In particular, p53’s role in maintaining genomic stability remains incompletely understood. It is well known that in the absence of normal p53 function, downstream effectors such as p21 are crippled and can no longer prevent aberrant cell cycling in response to DNA damage (4). However, this fact suggests that lack of p53 function is not directly responsible for genome instability, but instead that damaged DNA is allowed to inappropriately propagate through cell division if p53 function is absent (5). This mechanism would also suggest that increased cell cycling would produce more opportunities for DNA errors, and thus the absence of p53 function in this instance would allow cells with altered DNA to propagate rapidly, leading to an increased potential for producing oncogenic changes. However, many human cancers have low proliferation rates, yet still display genomic instability and aneuploidy (6). In addition, seminal studies have demonstrated that loss of TP53 has distinct functional consequences compared with TP53 missense mutations (7, 8), yet both types of alterations are found in human cancers. Thus, mechanisms of how genomic instability and aneuploidy arise may differ in cancer cells with homozygous loss of TP53 versus those with heterozygous missense mutations.In this study, we sought to elucidate the mechanism of genomic instability specifically associated with TP53 loss. We approached this via genome editing using the noncancerous human breast epithelial cell line, MCF10A, and comparing p53 null cells to isogenic cells harboring a common TP53 missense mutation, R248W. Relative to control and TP53 missense cell lines, we determined that p53 loss leads to increased genomic instability, which is associated with the presence of supernumerary centrosomes, a described mechanism of instability resulting in aneuploidy (9). Using an unbiased proteomic screen, we identified NDRG1 as differentially up-regulated in control and TP53 missense cell lines compared with TP53 null cells, but only under physiologic low-proliferation conditions. We show that forced expression of NDRG1 reduced abnormal centrosome numbers in MCF10A and HCT116 p53 null cells, whereas knockdown of N-Myc down-regulated gene 1 (NDRG1) by RNA interference (RNAi) in TP53 wild-type parental cells led to supernumerary centrosomes. Using proximity ligation assays, we found that NDRG1 associates with γ-tubulin, a key component of centrosomes, thus providing a mechanistic link between p53, NDRG1, and centrosome homeostasis. In silico analysis of multiple human tumor samples revealed that homozygous loss of TP53 is nearly mutually exclusive with NDRG1 overexpression, strongly corroborating our in vitro data to actual human cancers. Taken together, our results provide a previously unidentified model, suggesting that under conditions of physiologic low proliferation, p53 up-regulates NDRG1 expression, altering its interaction with γ-tubulin, thereby regulating centrosome homeostasis in a precise fashion. In cells with loss of p53, NDRG1 expression is not increased during cellular arrest and/or low proliferative states, allowing for supernumerary centrosome numbers, which results in genomic instability and aneuploidy.